Author: "Isabelle, Landrieu" - Searchworks@Jio Institute Digital Library Search Results

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144 results on '"Isabelle, Landrieu"'

1. Shaping the future of preclinical development of successful disease-modifying drugs against Alzheimer's disease: a systematic review of tau propagation models

Author: Neha Basheer, Luc Buee, Jean-Pierre Brion, Tomas Smolek, Muhammad Khalid Muhammadi, Jozef Hritz, Tomas Hromadka, Ilse Dewachter, Susanne Wegmann, Isabelle Landrieu, Petr Novak, Amritpal Mudher, and Norbert Zilka
Subjects: Tau protein, Propagation, Spreading, Aggregation, Neurofibrillary tangles, Animal models, Neurology. Diseases of the nervous system, RC346-429
Abstract: Abstract The transcellular propagation of the aberrantly modified protein tau along the functional brain network is a key hallmark of Alzheimer's disease and related tauopathies. Inoculation-based tau propagation models can recapitulate the stereotypical spread of tau and reproduce various types of tau inclusions linked to specific tauopathy, albeit with varying degrees of fidelity. With this systematic review, we underscore the significance of judicious selection and meticulous functional, biochemical, and biophysical characterization of various tau inocula. Furthermore, we highlight the necessity of choosing suitable animal models and inoculation sites, along with the critical need for validation of fibrillary pathology using confirmatory staining, to accurately recapitulate disease-specific inclusions. As a practical guide, we put forth a framework for establishing a benchmark of inoculation-based tau propagation models that holds promise for use in preclinical testing of disease-modifying drugs.
Published: 2024
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2. Dynamic interactions and Ca2+-binding modulate the holdase-type chaperone activity of S100B preventing tau aggregation and seeding

Author: Guilherme G. Moreira, François-Xavier Cantrelle, Andrea Quezada, Filipa S. Carvalho, Joana S. Cristóvão, Urmi Sengupta, Nicha Puangmalai, Ana P. Carapeto, Mário S. Rodrigues, Isabel Cardoso, Güenter Fritz, Federico Herrera, Rakez Kayed, Isabelle Landrieu, and Cláudio M. Gomes
Subjects: Science
Abstract: The calcium binding protein S100B is an abundantly expressed protein in the brain and has neuro-protective functions by inhibiting Aβ aggregation and metal ion toxicity. Here, the authors combine cell biology and biochemical experiments with chemical kinetics and NMR measurements and show that S100B protein is an extracellular Tau chaperone and further characterize the interactions between S100B and Tau.
Published: 2021
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3. Deciphering the Structure and Formation of Amyloids in Neurodegenerative Diseases With Chemical Biology Tools

Author: Isabelle Landrieu, Elian Dupré, Davy Sinnaeve, Léa El Hajjar, and Caroline Smet-Nocca
Subjects: amyloid fibril, aggregation, neurodegenerative diseases, protein semisynthesis, posttranslational modifications, native chemical ligation, Chemistry, QD1-999
Abstract: Protein aggregation into highly ordered, regularly repeated cross-β sheet structures called amyloid fibrils is closely associated to human disorders such as neurodegenerative diseases including Alzheimer’s and Parkinson’s diseases, or systemic diseases like type II diabetes. Yet, in some cases, such as the HET-s prion, amyloids have biological functions. High-resolution structures of amyloids fibrils from cryo-electron microscopy have very recently highlighted their ultrastructural organization and polymorphisms. However, the molecular mechanisms and the role of co-factors (posttranslational modifications, non-proteinaceous components and other proteins) acting on the fibril formation are still poorly understood. Whether amyloid fibrils play a toxic or protective role in the pathogenesis of neurodegenerative diseases remains to be elucidated. Furthermore, such aberrant protein-protein interactions challenge the search of small-molecule drugs or immunotherapy approaches targeting amyloid formation. In this review, we describe how chemical biology tools contribute to new insights on the mode of action of amyloidogenic proteins and peptides, defining their structural signature and aggregation pathways by capturing their molecular details and conformational heterogeneity. Challenging the imagination of scientists, this constantly expanding field provides crucial tools to unravel mechanistic detail of amyloid formation such as semisynthetic proteins and small-molecule sensors of conformational changes and/or aggregation. Protein engineering methods and bioorthogonal chemistry for the introduction of protein chemical modifications are additional fruitful strategies to tackle the challenge of understanding amyloid formation.
Published: 2022
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4. The elusive tau molecular structures: can we translate the recent breakthroughs into new targets for intervention?

Author: Yann Fichou, Youssra K. Al-Hilaly, François Devred, Caroline Smet-Nocca, Philipp O. Tsvetkov, Joke Verelst, Joris Winderickx, Nick Geukens, Eugeen Vanmechelen, Audrey Perrotin, Louise Serpell, Bernard J Hanseeuw, Miguel Medina, Luc Buée, and Isabelle Landrieu
Subjects: Tauopathies, Alzheimer’s disease, tau structure, tau aggregation, Alzheimer’s disease diagnosis, Amyloid, Neurology. Diseases of the nervous system, RC346-429
Abstract: Abstract Insights into tau molecular structures have advanced significantly in recent years. This field has been the subject of recent breakthroughs, including the first cryo-electron microscopy structures of tau filaments from Alzheimer’s and Pick’s disease inclusions, as well as the structure of the repeat regions of tau bound to microtubules. Tau structure covers various species as the tau protein itself takes many forms. We will here address a range of studies that help to define the many facets of tau protein structures and how they translate into pathogenic forms. New results shed light on previous data that need now to be revisited in order to up-date our knowledge of tau molecular structure. Finally, we explore how these data can contribute the important medical aspects of this research - diagnosis and therapeutics.
Published: 2019
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5. Phosphorylation and O-GlcNAcylation of the PHF-1 Epitope of Tau Protein Induce Local Conformational Changes of the C-Terminus and Modulate Tau Self-Assembly Into Fibrillar Aggregates

Author: François-Xavier Cantrelle, Anne Loyens, Xavier Trivelli, Oliver Reimann, Clément Despres, Neha S. Gandhi, Christian P. R. Hackenberger, Isabelle Landrieu, and Caroline Smet-Nocca
Subjects: Alzheimer’s disease, microtubule-associated protein tau, phosphorylation, O-GlcNAc glycosylation, protein aggregation, NMR spectroscopy, Neurosciences. Biological psychiatry. Neuropsychiatry, RC321-571
Abstract: Phosphorylation of the neuronal microtubule-associated Tau protein plays a critical role in the aggregation process leading to the formation of insoluble intraneuronal fibrils within Alzheimer’s disease (AD) brains. In recent years, other posttranslational modifications (PTMs) have been highlighted in the regulation of Tau (dys)functions. Among these PTMs, the O-β-linked N-acetylglucosaminylation (O-GlcNAcylation) modulates Tau phosphorylation and aggregation. We here focus on the role of the PHF-1 phospho-epitope of Tau C-terminal domain that is hyperphosphorylated in AD (at pS396/pS404) and encompasses S400 as the major O-GlcNAc site of Tau while two additional O-GlcNAc sites were found in the extreme C-terminus at S412 and S413. Using high resolution NMR spectroscopy, we showed that the O-GlcNAc glycosylation reduces phosphorylation of PHF-1 epitope by GSK3β alone or after priming by CDK2/cyclin A. Furthermore, investigations of the impact of PTMs on local conformation performed in small peptides highlight the role of S404 phosphorylation in inducing helical propensity in the region downstream pS404 that is exacerbated by other phosphorylations of PHF-1 epitope at S396 and S400, or O-GlcNAcylation of S400. Finally, the role of phosphorylation and O-GlcNAcylation of PHF-1 epitope was probed in in-vitro fibrillization assays in which O-GlcNAcylation slows down the rate of fibrillar assembly while GSK3β phosphorylation stimulates aggregation counteracting the effect of glycosylation.
Published: 2021
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6. Two Tau binding sites on tubulin revealed by thiol-disulfide exchanges

Author: Marlène Martinho, Diane Allegro, Isabelle Huvent, Charlotte Chabaud, Emilien Etienne, Hervé Kovacic, Bruno Guigliarelli, Vincent Peyrot, Isabelle Landrieu, Valérie Belle, and Pascale Barbier
Subjects: Thiol-disulfide Exchange, Label Release, Native Cysteine Residues, Spin Label, Co-sedimentation Assay, Medicine, Science
Abstract: Abstract Tau is a Microtubule-associated protein that induces and stabilizes the formation of the Microtubule cytoskeleton and plays an important role in neurodegenerative diseases. The Microtubules binding region of Tau has been determined for a long time but where and how Tau binds to its partner still remain a topic of debate. We used Site Directed Spin Labeling combined with EPR spectroscopy to monitor Tau upon binding to either Taxol-stabilized MTs or to αβ-tubulin when Tau is directly used as an inducer of MTs formation. Using maleimide-functionalized labels grafted on the two natural cysteine residues of Tau, we found in both cases that Tau remains highly flexible in these regions confirming the fuzziness of Tau:MTs complexes. More interestingly, using labels linked by a disulfide bridge, we evidenced for the first time thiol disulfide exchanges between αβ-tubulin or MTs and Tau. Additionally, Tau fragments having the two natural cysteines or variants containing only one of them were used to determine the role of each cysteine individually. The difference observed in the label release kinetics between preformed MTs or Tau-induced MTs, associated to a comparison of structural data, led us to propose two putative binding sites of Tau on αβ-tubulin.
Published: 2018
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7. Role of Tau as a Microtubule-Associated Protein: Structural and Functional Aspects

Author: Pascale Barbier, Orgeta Zejneli, Marlène Martinho, Alessia Lasorsa, Valérie Belle, Caroline Smet-Nocca, Philipp O. Tsvetkov, François Devred, and Isabelle Landrieu
Subjects: post-translational modifications, biophysical methods, Alzheimer’s disease, intrinsically disordered proteins, neurodegenerative diseases, Neurosciences. Biological psychiatry. Neuropsychiatry, RC321-571
Abstract: Microtubules (MTs) play a fundamental role in many vital processes such as cell division and neuronal activity. They are key structural and functional elements in axons, supporting neurite differentiation and growth, as well as transporting motor proteins along the axons, which use MTs as support tracks. Tau is a stabilizing MT associated protein, whose functions are mainly regulated by phosphorylation. A disruption of the MT network, which might be caused by Tau loss of function, is observed in a group of related diseases called tauopathies, which includes Alzheimer’s disease (AD). Tau is found hyperphosphorylated in AD, which might account for its loss of MT stabilizing capacity. Since destabilization of MTs after dissociation of Tau could contribute to toxicity in neurodegenerative diseases, a molecular understanding of this interaction and its regulation is essential.
Published: 2019
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8. Conformation and Affinity Modulations by Multiple Phosphorylation Occurring in the BIN1 SH3 Domain Binding Site of the Tau Protein Proline-Rich Region

Author: Alessia Lasorsa, Krishnendu Bera, Idir Malki, Elian Dupré, François-Xavier Cantrelle, Hamida Merzougui, Davy Sinnaeve, Xavier Hanoulle, Jozef Hritz, Isabelle Landrieu, Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), CHU Lille, Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Masaryk University [Brno] (MUNI), ANR-15-CE16-0002,BINALZ,Interaction entre BIN1 et Tau: consequences physiologiques et pathophysiologiques pour la maladie d'Alzheimer(2015), and ANR-11-LABX-0009,DISTALZ,Développement de stratégies innovantes pour une approche transdisciplinaire de la maladie d'Alzheime(2011)
Subjects: phosphorylation, protein-protein interactions, intrinsically disordered proteins, Alzheimer's disease, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Biochemistry, tau protein, molecular dynamics, nuclear magnetic resonance spectroscopy
Abstract: International audience; An increase in phosphorylation of the Tau protein is associated with Alzheimer's disease (AD) progression through unclear molecular mechanisms. In general, phosphorylation modifies the interaction of intrinsically disordered proteins, such as Tau, with other proteins; however, elucidating the structural basis of this regulation mechanism remains challenging. The bridging integrator-1 gene is an AD genetic determinant whose gene product, BIN1, directly interacts with Tau. The proline-rich motif recognized within a Tau(210-240) peptide by the SH3 domain of BIN1 (BIN1 SH3) is defined as 216PTPP219, and this interaction is modulated by phosphorylation. Phosphorylation of T217 within the Tau(210-240) peptide led to a 6-fold reduction in the affinity, while single phosphorylation at either T212, T231, or S235 had no effect on the interaction. Nonetheless, combined phosphorylation of T231 and S235 led to a 3-fold reduction in the affinity, although these phosphorylations are not within the BIN1 SH3-bound region of the Tau peptide. Using nuclear magnetic resonance (NMR) spectroscopy, these phosphorylations were shown to affect the local secondary structure and dynamics of the Tau(210-240) peptide. Models of the (un)phosphorylated peptides were obtained from molecular dynamics (MD) simulation validated by experimental data and showed compaction of the phosphorylated peptide due to increased salt bridge formation. This dynamic folding might indirectly impact the BIN1 SH3 binding by a decreased accessibility of the binding site. Regulation of the binding might thus not only be due to local electrostatic or steric effects from phosphorylation but also to the modification of the conformational properties of Tau.
Published: 2023
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9. A single domain antibody targeting Tau nucleation core inhibits Tau seeding

Author: Clément Danis, Elian Dupré, Orgeta Zejneli, Raphaëlle Caillierez, Alexis Arrial, Séverine Bégard, Anne Loyens, Justine Mortelecque, François-Xavier Cantrelle, Xavier Hanoulle, Jean-Christophe Rain, Morvane Colin, Luc Buée, and Isabelle Landrieu
Published: 2023
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10. Structural basis for oxygen degradation domain selectivity of the HIF prolyl hydroxylases

Author: Rasheduzzaman Chowdhury, Ivanhoe K. H. Leung, Ya-Min Tian, Martine I. Abboud, Wei Ge, Carmen Domene, François-Xavier Cantrelle, Isabelle Landrieu, Adam P. Hardy, Christopher W. Pugh, Peter J. Ratcliffe, Timothy D. W. Claridge, and Christopher J. Schofield
Subjects: Science
Abstract: The response to hypoxia involves multiple genes regulated by the hypoxia inducible transcription factors (HIFs), whose stability is regulated by prolyl hydroxylation. Here the authors provide a molecular basis for the substrate selectivity of the HIF prolyl hydroxylases that can be altered in erythrocytosis and cancer.
Published: 2016
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11. Structural Basis of Tau Interaction With BIN1 and Regulation by Tau Phosphorylation

Author: Alessia Lasorsa, Idir Malki, François-Xavier Cantrelle, Hamida Merzougui, Emmanuelle Boll, Jean-Charles Lambert, and Isabelle Landrieu
Subjects: protein-protein interaction, nuclear magnetic resonance spectroscopy, Alzheimer’s disease, bridging integrator-1 BIN1, SH3 domain, tau, Neurosciences. Biological psychiatry. Neuropsychiatry, RC321-571
Abstract: Bridging integrator-1 (BIN1) gene is associated with an increased risk to develop Alzheimer’s disease, a tauopathy characterized by intra-neuronal accumulation of phosphorylated Tau protein as paired helical filaments. Direct interaction of BIN1 and Tau proteins was demonstrated to be mediated through BIN1 SH3 C-terminal domain and Tau (210–240) peptide within Tau proline-rich domain. We previously showed that BIN1 SH3 interaction with Tau is decreased by phosphorylation within Tau proline-rich domain, of at least T231. In addition, the BIN1/Tau interaction is characterized by a dynamic equilibrium between a closed and open conformations of BIN1 isoform 1, involving an intramolecular interaction with its C-terminal BIN1 SH3 domain. However, the role of the BIN1/Tau interaction, and its potential dysregulation in Alzheimer’s disease, is not yet fully understood. Here we showed that within Tau (210–240) peptide, among the two proline-rich motifs potentially recognized by SH3 domains, only motif P216TPPTR221 is bound by BIN1 SH3. A structural model of the complex between BIN1 SH3 and Tau peptide (213–229), based on nuclear magnetic resonance spectroscopy data, revealed the molecular detail of the interaction. P216 and P219 within the proline-rich motif were in direct contact with the aromatic F588 and W562 of the BIN1 SH3 domain. The contact surface is extended through electrostatic interactions between the positively charged R221 and K224 residues of Tau peptide and those negatively charged of BIN1 SH3, corresponding to E556 and E557. We next investigated the impact of multiple Tau phosphorylations within Tau (210–240) on its interaction with BIN1 isoform 1. Tau (210–240) phosphorylated at four different sites (T212, T217, T231, and S235), contrary to unphosphorylated Tau, was unable to compete with the intramolecular interaction of BIN1 SH3 domain with its CLAP domain. In accordance, the affinity of BIN1 SH3 for phosphorylated Tau (210–240) peptide was reduced, with a five-fold increase in the dissociation constant, from a Kd of 44 to 256 μM. This study highlights the complexity of the regulation of BIN1 isoform 1 with Tau. As abnormal phosphorylation of Tau is linked to the pathology development, this regulation by phosphorylation might have important functional consequences.
Published: 2018
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12. Direct Crosstalk Between O-GlcNAcylation and Phosphorylation of Tau Protein Investigated by NMR Spectroscopy

Author: Gwendoline Bourré, François-Xavier Cantrelle, Amina Kamah, Béatrice Chambraud, Isabelle Landrieu, and Caroline Smet-Nocca
Subjects: Tau protein, phosphorylation, O-GlcNAcylation, crosstalk, NMR spectroscopy, Diseases of the endocrine glands. Clinical endocrinology, RC648-665
Abstract: The formation of intraneuronal fibrillar inclusions of tau protein is associated with several neurodegenerative diseases referred to as tauopathies including Alzheimer's disease (AD). A common feature of these pathologies is hyperphosphorylation of tau, the main component of fibrillar assemblies such as Paired Helical Filaments (PHFs). O-β-linked N-acetylglucosaminylation (O-GlcNAcylation) is another important posttranslational modification involved in regulation of tau pathophysiology. Among the benefits of O-GlcNAcylation, modulation of tau phosphorylation levels and inhibition of tau aggregation properties have been described while decreased O-GlcNAcylation could be involved in the raise of tau phosphorylation associated with AD. However, the molecular mechanisms at the basis of these observations remain to be defined. In this study, we identify by NMR spectroscopy O-GlcNAc sites in the longest isoform of tau and investigate the direct role of O-GlcNAcylation on tau phosphorylation and conversely, the role of phosphorylation on tau O-GlcNAcylation. We show here by a systematic examination of the quantitative modification patterns by NMR spectroscopy that O-GlcNAcylation does not modify phosphorylation of tau by the kinase activity of ERK2 or a rat brain extract while phosphorylation slightly increases tau O-GlcNAcylation by OGT. Our data suggest that indirect mechanisms act in the reciprocal regulation of tau phosphorylation and O-GlcNAcylation in vivo involving regulation of the enzymes responsible of phosphate and O-GlcNAc dynamics.
Published: 2018
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13. Dynamic interactions and Ca2+-binding modulate the holdase-type chaperone activity of S100B preventing tau aggregation and seeding

Author: Urmi Sengupta, Guilherme G. Moreira, Ana P. Carapeto, Filipa S. Carvalho, Andrea Quezada, Mário Rodrigues, Isabel Cardoso, Guenter Fritz, Nicha Puangmalai, Isabelle Landrieu, Federico Herrera, Rakez Kayed, Cláudio M. Gomes, Joana S. Cristóvão, François Xavier Cantrelle, Universidade de Lisboa = University of Lisbon (ULISBOA), Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), The University of Texas Medical Branch (UTMB), Universidade do Porto = University of Porto, Instituto de Ciências Biomédicas de Abel Salazar (ICBAS), University of Hohenheim, This work was funded by Fundação para a Ciência e Tecnologia (Portugal) through research grants PTDC/NEU-NMC/2138/2014 (to C.M.G.), PTDC/BIA-BQM/29963/2017 (F.S.C.), PTDC/MED-NEU/31417/2017 (to F.H.), and POCI-01-0145-FEDER-007274 (to I.C.), investigator grants CEECIND/00031/2017 (to A.P.C.) and IF/00094/2013/CP1173/CT0005 (to F.H.), PhD fellowship SFRH/BD/101171/2014 (to J.S.C.) and DFA/BD/6443/2020 (to G.G.M.), and center grants UIDB/04046/2020 and UID/MULTI/04046/2020 (to BioISI) and Norte-01-0145-FEDER-000008 (to IBMC/I3S)., Landrieu, Isabelle, Universidade de Lisboa (ULISBOA), and Universidade do Porto
Subjects: [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Science, Tau protein, General Physics and Astronomy, tau Proteins, S100 Calcium Binding Protein beta Subunit, Protein Aggregation, Pathological, Article, Biophysical Phenomena, General Biochemistry, Genetics and Molecular Biology, Cell Line, 03 medical and health sciences, 0302 clinical medicine, Microtubule, Biophysical chemistry, Calcium-binding protein, mental disorders, Humans, Protein folding, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Nuclear Magnetic Resonance, Biomolecular, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], 030304 developmental biology, 0303 health sciences, Multidisciplinary, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Neurodegenerative Diseases, General Chemistry, Kinetics, Proteostasis, Structural biology, Chaperone (protein), biology.protein, Biophysics, Protein Structural Elements, 030217 neurology & neurosurgery, Molecular Chaperones, Protein Binding, Binding domain
Abstract: The microtubule-associated protein tau is implicated in the formation of oligomers and fibrillar aggregates that evade proteostasis control and spread from cell-to-cell. Tau pathology is accompanied by sustained neuroinflammation and, while the release of alarmin mediators aggravates disease at late stages, early inflammatory responses encompass protective functions. This is the case of the Ca2+-binding S100B protein, an astrocytic alarmin which is augmented in AD and which has been recently implicated as a proteostasis regulator, acting over amyloid β aggregation. Here we report the activity of S100B as a suppressor of tau aggregation and seeding, operating at sub-stoichiometric conditions. We show that S100B interacts with tau in living cells even in microtubule-destabilizing conditions. Structural analysis revealed that tau undergoes dynamic interactions with S100B, in a Ca2+-dependent manner, notably with the aggregation prone repeat segments at the microtubule binding regions. This interaction involves contacts of tau with a cleft formed at the interface of the S100B dimer. Kinetic and mechanistic analysis revealed that S100B inhibits the aggregation of both full-length tau and of the microtubule binding domain, and that this proceeds through effects over primary and secondary nucleation, as confirmed by seeding assays and direct observation of S100B binding to tau oligomers and fibrils. In agreement with a role as an extracellular chaperone and its accumulation near tau positive inclusions, we show that S100B blocks proteopathic tau seeding. Together, our findings establish tau as a client of the S100B chaperone, providing evidence for neuro-protective functions of this inflammatory mediator across different tauopathies., The calcium binding protein S100B is an abundantly expressed protein in the brain and has neuro-protective functions by inhibiting Aβ aggregation and metal ion toxicity. Here, the authors combine cell biology and biochemical experiments with chemical kinetics and NMR measurements and show that S100B protein is an extracellular Tau chaperone and further characterize the interactions between S100B and Tau.
Published: 2021
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14. Tanycytes are degraded in Alzheimer’s Disease, disrupting the brain-to-blood efflux of Tau

Author: Florent Sauvé, Gaëtan Ternier, Julie Dewisme, Thibaut Lebouvier, Elian Dupré, Clément Danis, S. Rasika, Young-Bum Kim, Philippe Ciofi, Paolo Giacobini, Luc Buée, Isabelle Landrieu, Florence Pasquier, Claude-Alain Maurage, Ruben Nogueiras, Markus Schwaninger, and Vincent Prevot
Abstract: The accumulation of pathological Tau in the brain and cerebrospinal fluid (CSF) and its eventual increase in the blood are hallmarks of Alzheimer’s disease (AD). However, the mechanisms of Tau clearance from the brain to the periphery are not clear. We show here, using animal and cellular models as well as patient blood samples and post mortem brains, that hypothalamic tanycytes, whose cell bodies line the ventricular wall and send long processes to the underlying pituitary portal capillary bed, take up and transport Tau from the CSF and release it into these capillaries, whence it travels to the pituitary and eventually the systemic circulation. Specifically blocking tanycytic vesicular transport leads to an accumulation of exogenous fluorescent Tau in the CSF of mice. In AD and frontotemporal dementia, tanycytic morphology is altered, with a dramatic fragmentation of the secondary cytoskeleton in the former but not the latter, accounting for reduced CSF Tau clearance in AD. Both the implication of tanycytic degradation in the pathophysiology of a human disease and the evidence for the existence of a brain-to-blood tanycytic shuttle are unprecedented, and raise important questions regarding the role of tanycytes in physiological clearance mechanisms and the development of neurodegenerative disorders.
Published: 2022
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15. Design of Drug‐Like Protein–Protein Interaction Stabilizers Guided By Chelation‐Controlled Bioactive Conformation Stabilization

Author: Anders Gunnarsson, Gavin O'Mahony, Peter Brandt, Christian Ottmann, Francesco Bosica, Isabelle Landrieu, Sebastian A. Andrei, João Neves, Chemical Biology, ICMS Core, AstraZeneca, Gothenburg, Sweden, Technische Universiteit Eindhoven (TU/e), Biologie Structurale Intégrative (ERL 9002 - BSI ), Université de Lille, AstraZeneca, Eindhoven University of Technology [Eindhoven] (TU/e), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), University of Duisburg-Essen, Universität Duisburg-Essen = University of Duisburg-Essen [Essen], and Landrieu, Isabelle
Subjects: Drug, drug design, [CHIM.THER] Chemical Sciences/Medicinal Chemistry, [SDV]Life Sciences [q-bio], media_common.quotation_subject, Chemie, Molecular Conformation, Model system, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, 010402 general chemistry, 01 natural sciences, Catalysis, Protein–protein interaction, chemistry.chemical_compound, medicinal chemistry, Drug Discovery, protein–protein interaction stabilization, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Chelation, Glycosides, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], 14-3-3, Protein-protein interaction stabilization, Chelating Agents, media_common, PAINS, Biological Products, Natural product, 010405 organic chemistry, Chemistry, Organic Chemistry, Biomolecules (q-bio.BM), General Chemistry, Combinatorial chemistry, 0104 chemical sciences, chelates, Metal chelation, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Quantitative Biology - Biomolecules, 14-3-3 Proteins, Metals, FOS: Biological sciences, Protein Binding, Stabilizer (chemistry)
Abstract: International audience; The protein-protein interactions (PPIs) of 14-3-3 proteins are a model system for studying PPI stabilization. The complex natural product Fusicoccin A stabilizes many 14-3-3 PPIs but is not amenable for use in SAR studies, motivating the search for more drug-like chemical matter. However, drug-like 14-3-3 PPI stabilizers enabling such study have remained elusive. An X-ray crystal structure of a PPI in complex with an extremely low potency stabilizer uncovered an unexpected non-protein interacting, ligand-chelated Mg 2+ leading to the discovery of metal ion-dependent 14-3-3 PPI stabilization potency. This originates from a novel chelation-controlled bioactive conformation stabilization effect. Metal chelation has been associated with pan-assay interference compounds (PAINS) and frequent hitter behavior, but chelation can evidently also lead to true potency gains and find use as a medicinal chemistry strategy to guide compound optimization. To demonstrate this, we exploited the effect to design the first potent, selective and drug-like 14-3-3 PPI stabilizers.
Published: 2020
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16. Inhibition of Tau seeding by targeting Tau nucleation core within neurons with a single domain antibody fragment

Author: Clément Danis, Elian Dupré, Orgeta Zejneli, Raphaëlle Caillierez, Alexis Arrial, Séverine Bégard, Justine Mortelecque, Sabiha Eddarkaoui, Anne Loyens, François-Xavier Cantrelle, Xavier Hanoulle, Jean-Christophe Rain, Morvane Colin, Luc Buée, Isabelle Landrieu, Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Lille Neurosciences & Cognition - U 1172 (LilNCog), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Hybrigenics [Paris], Hybrigenics, ANR-18-CE44-0016,ToNIC,Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau(2018), and Landrieu, Isabelle
Subjects: Neurons, Pharmacology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], tau Proteins, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, Single-Domain Antibodies, Repressor Proteins, Disease Models, Animal, Mice, Tauopathies, Alzheimer Disease, Drug Discovery, Genetics, Animals, Molecular Medicine, [SDV.IMM.IMM] Life Sciences [q-bio]/Immunology/Immunotherapy, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Molecular Biology, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], ComputingMilieux_MISCELLANEOUS
Abstract: Tau proteins aggregate into filaments in brain cells in Alzheimer's disease and related disorders referred to as tauopathies. Here, we used fragments of camelid heavy-chain-only antibodies (VHHs or single domain antibody fragments) targeting Tau as immuno-modulators of its pathologic seeding. A VHH issued from the screen against Tau of a synthetic phage-display library of humanized VHHs was selected for its capacity to bind Tau microtubule-binding domain, composing the core of Tau fibrils. This parent VHH was optimized to improve its biochemical properties and to act in the intra-cellular compartment, resulting in VHH Z70. VHH Z70 precisely binds the PHF6 sequence, known for its nucleation capacity, as shown by the crystal structure of the complex. VHH Z70 was more efficient than the parent VHH to inhibit in vitro Tau aggregation in heparin-induced assays. Expression of VHH Z70 in a cellular model of Tau seeding also decreased the aggregation-reporting fluorescence signal. Finally, intra-cellular expression of VHH Z70 in the brain of an established tauopathy mouse seeding model demonstrated its capacity to mitigate accumulation of pathological Tau. VHH Z70, by targeting Tau inside brain neurons, where most of the pathological Tau resides, provides an immunological tool to target the intra-cellular compartment in tauopathies.
Published: 2022
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17. Comparative analysis of Erk phosphorylation suggests a mixed strategy for measuring phospho‐form distributions

Author: Sudhakaran Prabakaran, Robert A Everley, Isabelle Landrieu, Jean‐Michel Wieruszeski, Guy Lippens, Hanno Steen, and Jeremy Gunawardena
Subjects: Biology (General), QH301-705.5, Medicine (General), R5-920
Abstract: Abstract The functional impact of multisite protein phosphorylation can depend on both the numbers and the positions of phosphorylated sites—the global pattern of phosphorylation or ‘phospho‐form’—giving biological systems profound capabilities for dynamic information processing. A central problem in quantitative systems biology, therefore, is to measure the ‘phospho‐form distribution’: the relative amount of each of the 2n phospho‐forms of a protein with n‐phosphorylation sites. We compared four potential methods—western blots with phospho‐specific antibodies, peptide‐based liquid chromatography (LC) and mass spectrometry (MS; pepMS), protein‐based LC/MS (proMS) and nuclear magnetic resonance spectroscopy (NMR)—on differentially phosphorylated samples of the well‐studied mitogen‐activated protein kinase Erk2, with two phosphorylation sites. The MS methods were quantitatively consistent with each other and with NMR to within 10%, but western blots, while highly sensitive, showed significant discrepancies with MS. NMR also uncovered two additional phosphorylations, for which a combination of pepMS and proMS yielded an estimate of the 16‐member phospho‐form distribution. This combined MS strategy provides an optimal mixture of accuracy and coverage for quantifying distributions, but positional isomers remain a challenging problem.
Published: 2011
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18. Inhibition of Tau seeding by targeting Tau nucleation core within neurons with a single domain antibody fragment

Author: Séverine Bégard, Alexis Arrial, Danis C, Raphaëlle Caillierez, E. Dupre, Xavier Hanoulle, Isabelle Landrieu, Zejneli O, Mortelecque J, Anne Loyens, Francois-Xavier Cantrelle, Morvane Colin, Luc Buée L, Jean-Christophe Rain, Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Lille Neurosciences & Cognition - U 1172 (LilNCog (ex-JPARC)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Hybrigenics [Paris], Hybrigenics, The NMR facilities were funded by the Nord Region Council, CNRS, Institut Pasteur de Lille, European Union (FEDER), French Research Ministry and Univ. Lille. We acknowledge support from TGE RMN THC (FR-3050, France). This study was supported by the LabEx (Laboratory of Excellence) DISTALZ (Development of Innovative Strategies for a Transdisciplinary approach to Alzheimer’s disease ANR-11-LABX-01), by EU project AgedBrainSYSBIO (Grant Agreement N° 305299), by I-site ULNE (project TUNABLE) and by ANR (project ToNIC). Our laboratories are also supported by LiCEND (Lille Centre of Excellence in Neurodegenerative Disorders), Inserm, Métropole Européenne de Lille, Univ. Lille and FEDER., ANR-11-LABX-0001,ACTION,Systèmes intelligents intégrés au cœur de la matière(2011), ANR-18-CE44-0016,ToNIC,Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau(2018), European Project: 305299,EC:FP7:HEALTH,FP7-HEALTH-2012-INNOVATION-1,AGEDBRAINSYSBIO(2013), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Lille Neurosciences & Cognition - U 1172 (LilNCog), Landrieu, Isabelle, Laboratoires d'excellence - Systèmes intelligents intégrés au cœur de la matière - - ACTION2011 - ANR-11-LABX-0001 - LABX - VALID, APPEL À PROJETS GÉNÉRIQUE 2018 - Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau - - ToNIC2018 - ANR-18-CE44-0016 - AAPG2018 - VALID, and SYSTEMS BIOLOGY OF PATHWAYS INVOLVING BRAIN AGEING - AGEDBRAINSYSBIO - - EC:FP7:HEALTH2013-01-01 - 2016-12-31 - 305299 - VALID
Subjects: medicine.medical_treatment, [SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, [SDV.BBM.BP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, VHH, Fibril, protein-protein interaction, 03 medical and health sciences, 0302 clinical medicine, mental disorders, medicine, Tau protein, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], 030304 developmental biology, nuclear magnetic resonance spectroscopy, 0303 health sciences, biology, Chemistry, tauopathies, [SDV.NEU.NB] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, aggregation, Immunotherapy, medicine.disease, In vitro, nanobodies, 3. Good health, Cell biology, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, Single-domain antibody, biology.protein, Tauopathy, immunotherapy, Antibody, Cellular model, 030217 neurology & neurosurgery, Intracellular, seeding
Abstract: Tau proteins aggregate into filaments in brain cells in Alzheimer’s disease and related disorders referred to as tauopathies. Here, we used fragments of camelid heavy-chain-only antibodies (VHHs or single domain antibody fragments) targeting Tau as immuno-modulators of its pathologic seeding. A VHH issued from the screen against Tau of a synthetic phage-display library of humanized VHHs was selected for its capacity to bind Tau microtubule-binding domain, composing the core of Tau fibrils. This lead VHH was optimized to improve its biochemical properties and to act in the intracellular compartment, resulting in VHH Z70. VHH Z70 was more efficient than the lead to inhibit in vitro Tau aggregation in heparin-induced assays. Expression of VHH Z70 in a cellular model of Tau seeding also decreased the fluorescence-reported aggregation. Finally, intracellular expression of VHH Z70 in the brain of an established tauopathy mouse seeding model demonstrated its capacity to mitigate accumulation of pathological Tau. VHH Z70, by targeting Tau inside brain neurons, where most of the pathological Tau resides, provides a new tool to explore the optimal strategies of immunotherapy in tauopathies.
Published: 2021

19. NMR spectroscopy of the main protease of SARS‐CoV‐2 and fragment‐based screening identify three protein hotspots and an antiviral fragment

Author: Florence Leroux, Julie Charton, François-Xavier Cantrelle, Xavier Hanoulle, Sandrine Belouzard, Danai Moschidi, Isabelle Landrieu, Lucile Brier, Jean Dubuisson, Valérie Landry, Emmanuelle Boll, Benoit Deprez, Frédérique Dewitte, Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Biologie Structurale Intégrative (ERL 9002 - BSI ), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Médicaments et molécules pour agir sur les Systèmes Vivants - U 1177 (M2SV), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS), Projet 3CLPRO-SCREEN-NMR (I-SITE Université Lille Nord-Europe)Institut Pasteur de Lille, HANOULLE, Xavier, Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, and Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)
Subjects: Protein Conformation, medicine.medical_treatment, Drug Evaluation, Preclinical, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, 01 natural sciences, Protein structure, Chlorocebus aethiops, Coronavirus 3C Proteases, Research Articles, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, 0303 health sciences, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Drug discovery, Chemistry, fragment screening, Nuclear magnetic resonance spectroscopy, General Medicine, 3. Good health, Protein Structure | Hot Paper, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, Research Article, Stereochemistry, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], [CHIM.THER] Chemical Sciences/Medicinal Chemistry, Allosteric regulation, Context (language use), Microbial Sensitivity Tests, Cysteine Proteinase Inhibitors, 010402 general chemistry, Antiviral Agents, Catalysis, drug discovery, Small Molecule Libraries, 03 medical and health sciences, NMR spectroscopy, medicine, Animals, viruses, Binding site, protein structure, Nuclear Magnetic Resonance, Biomolecular, Vero Cells, 030304 developmental biology, Protease, Binding Sites, SARS-CoV-2, 010405 organic chemistry, Active site, General Chemistry, 0104 chemical sciences, biology.protein, Protein Multimerization
Abstract: The main protease (3CLp) of the SARS‐CoV‐2, the causative agent for the COVID‐19 pandemic, is one of the main targets for drug development. To be active, 3CLp relies on a complex interplay between dimerization, active site flexibility, and allosteric regulation. The deciphering of these mechanisms is a crucial step to enable the search for inhibitors. In this context, using NMR spectroscopy, we studied the conformation of dimeric 3CLp from the SARS‐CoV‐2 and monitored ligand binding, based on NMR signal assignments. We performed a fragment‐based screening that led to the identification of 38 fragment hits. Their binding sites showed three hotspots on 3CLp, two in the substrate binding pocket and one at the dimer interface. F01 is a non‐covalent inhibitor of the 3CLp and has antiviral activity in SARS‐CoV‐2 infected cells. This study sheds light on the complex structure‐function relationships of 3CLp and constitutes a strong basis to assist in developing potent 3CLp inhibitors., We report the liquid‐sate NMR spectroscopy analysis of the dimeric SARS‐CoV‐2 main protease (3CLp), including its backbone assignments, to study its complex conformational regulation. Using fragment‐based NMR screening, we highlighted three hotspots on the protein, two in the substrate binding pocket and one at the dimer interface, and we identified a non‐covalent reversible inhibitor of 3CLp that has antiviral activity in infected cells.
Published: 2021
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20. Set-up and screening of a fragment library targeting the 14-3-3 protein interface

Author: Dimitrios Tzalis, Christian Ottmann, Laura Levy, Domenico Lentini Santo, Stanimira Hristeva, Isabelle Landrieu, João Neves, Tomas Obsil, Dario Valenti, François Xavier Cantrelle, Xavier Hanoulle, Taros Chemicals GmbH & Co. KG, Technische Universiteit Eindhoven (TU/e), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Charles University [Prague] (CU), Universität Duisburg-Essen [Essen], Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Charles University [Prague], Chemical Biology, Université de Lille-Centre National de la Recherche Scientifique (CNRS), and Universität Duisburg-Essen = University of Duisburg-Essen [Essen]
Subjects: Pharmacology, Gene isoform, 010405 organic chemistry, Chemistry, Organic Chemistry, Chemie, Pharmaceutical Science, Signal transducing adaptor protein, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Nuclear magnetic resonance spectroscopy, Computational biology, 01 natural sciences, Biochemistry, 0104 chemical sciences, Selective modulation, 010404 medicinal & biomolecular chemistry, Fragment (logic), Drug Discovery, Molecular Medicine, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 14-3-3 protein
Abstract: Protein-protein interactions (PPIs) are at the core of regulation mechanisms in biological systems and consequently became an attractive target for therapeutic intervention. PPIs involving the adapter protein 14-3-3 are representative examples given the broad range of partner proteins forming a complex with one of its seven human isoforms. Given the challenges represented by the nature of these interactions, fragment-based approaches offer a valid alternative for the development of PPI modulators. After having assembled a fragment set tailored on PPIs' modulation, we started a screening campaign on the sigma isoform of 14-3-3 adapter proteins. Through the use of both mono-and bi-dimensional nuclear magnetic resonance spectroscopy measurements, coupled with differential scanning fluorimetry, three fragment hits were identified. These molecules bind the protein at two different regions distant from the usual binding groove highlighting new possibilities for selective modulation of 14-3-3 complexes.
Published: 2019
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21. Phosphorylated full-length Tau interacts with 14-3-3 proteins via two short phosphorylated sequences, each occupying a binding groove of 14-3-3 dimer

Author: João Filipe Neves, François Xavier Cantrelle, Tomas Obsil, Isabelle Landrieu, Gavin O'Mahony, Xavier Hanoulle, Olivia Petrvalska, Francesco Bosica, Hamida Merzougui, Chemical Biology, Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Charles University [Prague] (CU), AstraZeneca, European Project: 675179,H2020,H2020-MSCA-ITN-2015,TASPPI(2016), Landrieu, Isabelle, and Targeted small-molecule Stabilisation of Protein-Protein Interactions - TASPPI - - H20202016-02-01 - 2020-01-31 - 675179 - VALID
Subjects: 0301 basic medicine, Magnetic Resonance Spectroscopy, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein Conformation, Dimer, Tau protein, tau Proteins, protein–protein interactions, Biochemistry, Epitope, Protein–protein interaction, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, NMR spectroscopy, Alzheimer Disease, Serine, Humans, Phosphorylation, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Molecular Biology, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Binding Sites, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Drug discovery, 14-3-3 proteins, Cell Biology, Nuclear magnetic resonance spectroscopy, Surface Plasmon Resonance, Alzheimer's disease, Cyclic AMP-Dependent Protein Kinases, 030104 developmental biology, Monomer, Protein-Protein Interactions, 030220 oncology & carcinogenesis, Exoribonucleases, Mutation, Biophysics, biology.protein, Protein Multimerization, Alzheimer’s disease, analytical ultracentrifugation, Protein Binding
Abstract: Protein-protein interactions (PPIs) remain poorly explored targets for the treatment of Alzheimer’s disease (AD). The interaction of 14-3-3 proteins with Tau was shown to have detrimental effects on neuronal cells and to be linked to Tau pathology. This PPI is therefore seen as a potential target for AD. When Tau is phosphorylated by PKA (Tau-PKA), two 14-3-3 binding epitopes are generated, surrounding the phosphorylated serines 214 and 324 of Tau. The crystal structures of 14-3-3 in complex with peptides surrounding these Tau phosphosites show that both these motifs are anchored in the amphipathic binding groove of 14-3-3. However, in the absence of structural data with the full-length Tau protein, the stoichiometry of the complex or the interface and affinity of the partners, are still unclear. In this work, we addressed these points, using a broad range of biophysical techniques. The interaction of the long disordered Tau-PKA protein with 14-3-3σ is restricted to two short sequences, containing phosphorylated serines, which bind in the amphipathic binding groove of 14-3-3. Phosphorylation of Tau is fundamental for the formation of this stable complex, and the affinity of the Tau-PKA/14-3-3 interaction is in the 1-10 micromolar range. Each monomer of the 14-3-3σ dimer binds one of the two different phosphorylated peptides of Tau-PKA, suggesting a 14-3-3/Tau-PKA stoichiometry of 2:1, confirmed by analytical ultracentrifugation. These results contribute to a better understanding of this PPI and provide useful insights for drug discovery projects aiming at the inhibition of this interaction.
Published: 2021
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22. NMR Meets Tau: Insights into Its Function and Pathology

Author: Guy Lippens, Isabelle Landrieu, Caroline Smet, Isabelle Huvent, Neha S. Gandhi, Benoît Gigant, Clément Despres, Haoling Qi, and Juan Lopez
Subjects: Tau, NMR spectroscopy, intrinsically disordered protein, tubulin, aggregation, phosphorylation, protein/protein interactions, Microbiology, QR1-502
Abstract: In this review, we focus on what we have learned from Nuclear Magnetic Resonance (NMR) studies on the neuronal microtubule-associated protein Tau. We consider both the mechanistic details of Tau: the tubulin relationship and its aggregation process. Phosphorylation of Tau is intimately linked to both aspects. NMR spectroscopy has depicted accurate phosphorylation patterns by different kinases, and its non-destructive character has allowed functional assays with the same samples. Finally, we will discuss other post-translational modifications of Tau and its interaction with other cellular factors in relationship to its (dys)function.
Published: 2016
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23. 1H, 13C, and 15N chemical shift assignment of human PACSIN1/syndapin I SH3 domain in solution

Author: Isabelle Landrieu, Geraldine Levy, Davy Sinnaeve, Matthieu Hirel, Francois-Xavier Cantrelle, Emmanuelle Boll, Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Biologie Structurale Intégrative (ERL 9002 - BSI ), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), The research is supported by funding from the Métropole Européenne de Lille, through the PUSHUP project. We acknowledge LabEx (Laboratory of Excellence) for financial support on the scope of the DISTALZ consortium (ANR, ANR-11-LABX-009). The NMR facilities were funded by the Nord Region Council, CNRS, Institut Pasteur de Lille, the European Community (ERDF), the French Ministry of Research and the Université de Lille and by the CTRL CPER cofunded by the European Union with the European Regional Development Fund (ERDF), by the Hauts-de-France Regional Council (Contract No 17003781), Métropole Européenne de Lille (Contract No 2016_ESR_05), and French State (Contract No 2017-R3-CTRL-Phase1). We acknowledge support for the NMR facilities from TGE RMN THC (CNRS, FR-3050) and FRABio (Univ. Lille, CNRS, FR-3688)., ANR-11-LABX-0009,DISTALZ,Développement de stratégies innovantes pour une approche transdisciplinaire de la maladie d'Alzheime(2011), Mortelecque, Justine, and Laboratoires d'excellence - Développement de stratégies innovantes pour une approche transdisciplinaire de la maladie d'Alzheime - - DISTALZ2011 - ANR-11-LABX-0009 - LABX - VALID
Subjects: 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Stereochemistry, Chemistry, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], PACSIN1, Chemical shift, 030303 biophysics, IN1, Endocytosis, Biochemistry, NMR resonance assignment, SH3 domain, Protein–protein interaction, protein-protein interaction, 03 medical and health sciences, Structural Biology, Domain (ring theory), Synaptic vesicle recycling, Protein secondary structure, Heteronuclear single quantum coherence spectroscopy, 030304 developmental biology
Abstract: International audience; Human neuron-specific PACSIN1 plays a key role in synaptic vesicle recycling and endocytosis, as well as reorganization of the microtubule dynamics to maintain axonal plasticity. PACSIN1 contains a highly conserved C-terminal SH3 domain and an F-bar domain at its N-terminus. Due to its remarkable interaction network, PACSIN1 plays a central role in key neuronal functions. Here, we present a robust backbone and side-chain assignment of PACSIN1 SH3 domain based on 2D [1H,15N] HSQC or HMQC, and 3D BEST-HNCO, -HNCACB, -HN(CO)CACB, -HN(CA)CO, and standard (H)CC(CO)NH, HN(CA)NNH, HN(COCA)NH, HBHANNH, HNHA, HBHA(CO)NH, H(CC)(CO)NH, HCCH-TOCSY, that covers 96% for all 13CO, 13Cα and 13Cβ, 28% of 13Cγδε, and 95% of 1HN and 15N chemical shifts. Modelling based on sequence homology with a known related structure, and chemical shift-based secondary structure predictions, identified the presence of five β-strands linked by flexible loops. Taken together, these results open up new avenues to investigate and develop new therapeutic strategies.
Published: 2020
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24. Selectivity via Cooperativity : Preferential Stabilization of the p65/14-3-3 Interaction with Semisynthetic Natural Products

Author: João Neves, Christian Ottmann, Sonja Srdanovic, M. Wolter, Nobuo Kato, Luc Brunsveld, Pim J. de Vink, Isabelle Landrieu, Yusuke Higuchi, Andrew J. Wilson, Eindhoven University of Technology [Eindhoven] (TU/e), Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 (RID-AGE), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Biologie Structurale Intégrative (ERL 9002 - BSI ), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), University of Leeds, Osaka University [Osaka], University of Duisburg-Essen, This research is supported by funding from the European Union through the TASPPI project (H2020-MSCA-ITN-2015, grant number 675179) and by the LabEx (Laboratory of Excellence) DISTALZ (ANR, ANR-11-LABX-009) and through The Netherlands Organization for Scientific Research (NWO) via VICI grant 016.150.366 and via Gravity Program 024.001.035. The NMR facilities were funded by the Nord Region Council, CNRS, Institut Pasteur de Lille, the European Community (ERDF), the French Ministry of Research, and the University of Lille. We acknowledge support for the NMR facilities from TGE RMN THC (CNRS, FR-3050) and FRABio (Univ. Lille, CNRS, FR-3688). This work was supported by the EPSRC (EP/N013573/1) and The Wellcome Trust (097827/Z/11/A)., ANR-11-LABX-0009,DISTALZ,Développement de stratégies innovantes pour une approche transdisciplinaire de la maladie d'Alzheime(2011), European Project: 675179,H2020,H2020-MSCA-ITN-2015,TASPPI(2016), Université de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Chemical Biology, ICMS Core, and Universität Duisburg-Essen = University of Duisburg-Essen [Essen]
Subjects: Scaffold protein, Models, Molecular, Protein subunit, Chemie, Cooperativity, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, 010402 general chemistry, 01 natural sciences, Biochemistry, Catalysis, Article, Protein–protein interaction, chemistry.chemical_compound, Colloid and Surface Chemistry, Humans, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], ComputingMilieux_MISCELLANEOUS, Biological Products, Natural product, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Drug discovery, NF-kappa B, Cooperative binding, General Chemistry, 0104 chemical sciences, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], chemistry, 14-3-3 Proteins, Biophysics, Stabilizer (chemistry), Protein Binding
Abstract: International audience; Natural compounds are an important class of potent drug molecules including some retrospectively found to act as stabilizers of protein–protein interactions (PPIs). However, the design of synthetic PPI stabilizers remains an understudied approach. To date, there are limited examples where cooperativity has been utilized to guide the optimization of a PPI stabilizer. The 14-3-3 scaffold proteins provide an excellent platform to explore PPI stabilization because these proteins mediate several hundred PPIs, and a class of natural compounds, the fusicoccanes, are known to stabilize a subset of 14-3-3 protein interactions. 14-3-3 has been reported to negatively regulate the p65 subunit of the NF-κB transcription factor, which qualifies this protein complex as a potential target for drug discovery to control cell proliferation. Here, we report the high-resolution crystal structures of two 14-3-3 binding motifs of p65 in complex with 14-3-3. A semisynthetic natural product derivative, DP-005, binds to an interface pocket of the p65/14-3-3 complex and concomitantly stabilizes it. Cooperativity analyses of this interaction, and other disease relevant 14-3-3-PPIs, demonstrated selectivity of DP-005 for the p65/14-3-3 complex. The adaptation of a cooperative binding model provided a general approach to characterize stabilization and to assay for selectivity of PPI stabilizers.
Published: 2020
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25. Modulators of 14-3-3 protein-protein interactions

Author: Ylenia Cau, Loes M. Stevers, Jeremy Martin Davis, Eline Sijbesma, Maurizio Botta, Carol MacKintosh, Andrew J. Wilson, Christian Ottmann, Isabelle Landrieu, Jan Eickhoff, Richard G. Doveston, Luc Brunsveld, GGavin O'Mahony, Tomas Obsil, Anna Karawajczyk, Michael M. Hann, CNRS, Université de Lille, Eindhoven University of Technology [Eindhoven] [TU/e], Università degli Studi di Siena = University of Siena [UNISI], University of Dundee, Charles University [Prague] [CU], Unité de Glycobiologie Structurale et Fonctionnelle (UGSF) - UMR 8576, University of Leeds, GlaxoSmithKline [Stevenage, UK] [GSK], AstraZeneca, Chemical Biology, Eindhoven University of Technology [Eindhoven] (TU/e), Dipartimento Farmaco Chimico Tecnologico, Università degli Studi di Siena, College of Life Sciences, MRC Protein Phosphorylation Unit, University of Pragues, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Universität Duisburg-Essen [Essen], UCB Pharma Slough, AstraZeneca (AstraZeneca), Technische Universiteit Eindhoven (TU/e), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Università degli Studi di Siena = University of Siena (UNISI), Charles University [Prague] (CU), Université de Lille-Centre National de la Recherche Scientifique (CNRS), School of Chemistry [Leeds], Universität Duisburg-Essen = University of Duisburg-Essen [Essen], and Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Nanotechnology, Computational biology, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Protein–protein interaction, 03 medical and health sciences, Drug Discovery, Animals, Humans, [CHIM]Chemical Sciences, Peptides and proteins, Monomers, Crystal structure, Inhibitors, Inhibition, 14-3-3 protein, ComputingMilieux_MISCELLANEOUS, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Protein Stability, Drug Discovery3003 Pharmaceutical Science, Small molecule, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030104 developmental biology, 14-3-3 Proteins, Molecular Medicine, Perspective, Medical science, Biologie, Protein Binding
Abstract: International audience; Direct interactions between proteins are essential for the regulation of their functions in biological pathways. Targeting the complex network of protein–protein interactions (PPIs) has now been widely recognized as an attractive means to therapeutically intervene in disease states. Even though this is a challenging endeavor and PPIs have long been regarded as “undruggable” targets, the last two decades have seen an increasing number of successful examples of PPI modulators, resulting in growing interest in this field. PPI modulation requires novel approaches and the integrated efforts of multiple disciplines to be a fruitful strategy. This perspective focuses on the hub-protein 14-3-3, which has several hundred identified protein interaction partners, and is therefore involved in a wide range of cellular processes and diseases. Here, we aim to provide an integrated overview of the approaches explored for the modulation of 14-3-3 PPIs and review the examples resulting from these efforts in both inhibiting and stabilizing specific 14-3-3 protein complexes by small molecules, peptide mimetics, and natural products.
Published: 2018
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26. The O-β-linked N-acetylglucosaminylation of the Lamin B receptor and its impact on DNA binding and phosphorylation

Author: Caroline Smet-Nocca, Eleni Nikolakaki, Isabelle Landrieu, Adeline Page, François-Xavier Cantrelle, Thomas Giannakouros, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Centre National de la Recherche Scientifique (CNRS), Aristotelian University of Thessaloniki, Aristotle University of Thessaloniki, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Turkeys, Glycosylation, Proto-Oncogene Proteins c-akt, [SDV]Life Sciences [q-bio], Biophysics, Receptors, Cytoplasmic and Nuclear, Lamin B receptor, N-Acetylglucosaminyltransferases, Biochemistry, Acetylglucosamine, 03 medical and health sciences, CDC2 Protein Kinase, Animals, Humans, Inner membrane, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Protein phosphorylation, Amino Acid Sequence, Phosphorylation, Nuclear protein, Molecular Biology, Integral membrane protein, ComputingMilieux_MISCELLANEOUS, Binding Sites, Sequence Homology, Amino Acid, Chemistry, DNA, Rats, Chromatin, Cell biology, 030104 developmental biology, Peptides, Protein Binding
Abstract: Lamin B Receptor (LBR) is an integral protein of the interphase inner nuclear membrane that is implicated in chromatin anchorage to the nuclear envelope. Phosphorylation of a stretch of arginine-serine (RS) dipeptides in the amino-terminal nucleoplasmic domain of LBR regulates the interactions of the receptor with other nuclear proteins, DNA and RNA and thus modulates tethering of heterochromatin to the nuclear envelope. While phosphorylation has been extensively studied, very little is known about other post-translational modifications of the protein. There is only one report on the O-β-linked N-acetyl-glucosaminylation (O-GlcNAcylation) of a serine residue downstream of the RS domain of rat LBR. In the present study we identify additional O-GlcNAcylation sites by using as substrates of O-β-N-acetylglucosaminyltransferase (OGT) a set of peptides containing the entire LBR RS domain or parts of it as well as flanking sequences. The in vitro activity of OGT was assessed by tandem mass spectrometry and NMR spectroscopy. Furthermore, we provide evidence that O-GlcNAcylation hampers DNA binding while it marginally affects RS domain phosphorylation mediated by SRPK1, Akt2 and cdk1 kinases. General significance Our methodology providing a quantitative description of O-GlcNAc patterns based on a combination of mass spectrometry and high resolution NMR spectroscopy on short peptide substrates allows subsequent functional analyses. Hence, our approach is of general interest to a wide audience of biologists aiming at deciphering the functional role of O-GlcNAc glycosylation and its crosstalk with phosphorylation.
Published: 2018
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27. Molecular implication of PP2A and Pin1 in the Alzheimer's disease specific hyperphosphorylation of Tau.

Author: Isabelle Landrieu, Caroline Smet-Nocca, Laziza Amniai, Justin Vijay Louis, Jean-Michel Wieruszeski, Jozef Goris, Veerle Janssens, and Guy Lippens
Subjects: Medicine, Science
Abstract: Tau phosphorylation and dephosphorylation regulate in a poorly understood manner its physiological role of microtubule stabilization, and equally its integration in Alzheimer disease (AD) related fibrils. A specific phospho-pattern will result from the balance between kinases and phosphatases. The heterotrimeric Protein Phosphatase type 2A encompassing regulatory subunit PR55/Bα (PP2A(T55α)) is a major Tau phosphatase in vivo, which contributes to its final phosphorylation state. We use NMR spectroscopy to determine the dephosphorylation rates of phospho-Tau by this major brain phosphatase, and present site-specific and kinetic data for the individual sites including the pS202/pT205 AT8 and pT231 AT180 phospho-epitopes.We demonstrate the importance of the PR55/Bα regulatory subunit of PP2A within this enzymatic process, and show that, unexpectedly, phosphorylation at the pT231 AT180 site negatively interferes with the dephosphorylation of the pS202/pT205 AT8 site. This inhibitory effect can be released by the phosphorylation dependent prolyl cis/trans isomerase Pin1. Because the stimulatory effect is lost with the dimeric PP2A core enzyme (PP2A(D)) or with a phospho-Tau T231A mutant, we propose that Pin1 regulates the interaction between the PR55/Bα subunit and the AT180 phospho-epitope on Tau.Our results show that phosphorylation of T231 (AT180) can negatively influence the dephosphorylation of the pS202/pT205 AT8 epitope, even without an altered PP2A pool. Thus, a priming dephosphorylation of pT231 AT180 is required for efficient PP2A(T55α)-mediated dephosphorylation of pS202/pT205 AT8. The sophisticated interplay between priming mechanisms reported for certain Tau kinases and the one described here for Tau phosphatase PP2A(T55α) may contribute to the hyperphosphorylation of Tau observed in AD neurons.
Published: 2011
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28. Zinc Binding to Tau Influences Aggregation Kineticsand Oligomer Distribution

Author: Isabelle Landrieu, Mário Rodrigues, Carlos Cordeiro, Guilherme G. Moreira, Joana S. Cristóvão, Vukosava M Torres, Cláudio M. Gomes, Ana P. Carapeto, Universidade de Lisboa (ULISBOA), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Universidade de Lisboa = University of Lisbon (ULISBOA), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), and Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: Circular dichroism, spectroscopy, Amyloid, Protein Conformation, Tau protein, chemistry.chemical_element, tau Proteins, Zinc, Protein aggregation, Oligomer, Article, Catalysis, protein aggregation, Inorganic Chemistry, Protein Aggregates, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Dynamic light scattering, Alzheimer Disease, Spectroscopy, Fourier Transform Infrared, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Benzothiazoles, Physical and Theoretical Chemistry, Molecular Biology, Conformational isomerism, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, mass spectrometry, 0303 health sciences, calcium, biology, Chemistry, Circular Dichroism, Organic Chemistry, zinc, neurodegeneration, amyloid, General Medicine, intrinsically disordered protein, Computer Science Applications, Kinetics, stomatognathic diseases, Biophysics, biology.protein, Tau, human activities, 030217 neurology & neurosurgery
Abstract: Metal ions are well known modulators of protein aggregation and are key players in Alzheimer&rsquo, s Disease, being found to be associated to pathologic protein deposits in diseased brains. Therefore, understanding how metals influence amyloid aggregation is critical in establishing molecular mechanisms that underlie disease onset and progression. Here, we report data on the interaction of full-length human Tau protein with calcium and zinc ions, evidencing that Tau self-assembly is differently regulated, depending on the type of bound metal ion. We established that Tau binds 4 Zn2+ and 1 Ca2+ per monomer while using native mass spectrometry analysis, without inducing order or substantial conformational changes in the intrinsically disordered Tau, as determined by structural analysis using circular dichroism and Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR) spectroscopies. However, Tau aggregation is found to proceed differently in the calcium- and -zinc bound forms. While the rate of aggregation, as determined from thioflavin-T (ThT) fluorescence kinetics, is highly increased in both cases, the reaction proceeds via different mechanisms, as evidenced by the absence of the lag phase in the reaction of zinc-bound Tau. Monitoring Tau aggregation using native mass spectrometry indeed evidenced a distinct distribution of Tau conformers along the reaction, as confirmed by dynamic light scattering analysis. We propose that such differences arise from zinc binding at distinct locations within the Tau sequence that prompt both the rapid formation of seeding oligomers through interactions at high affinity sites within the repeat domains, as well as amorphous aggregation, through low affinity interactions with residues elsewhere in the sequence, including at the fuzzy coat domain.
Published: 2019
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29. Single Domain Antibody Fragments as New Tools for the Detection of Neuronal Tau Protein in Cells and in Mice Studies

Author: Clément Danis, Morvane Colin, Isabelle Landrieu, Elian Dupre, Jean-Christophe Rain, François-Xavier Cantrelle, Mégane Homa, Hamida Merzougui, Alexis Arrial, Luc Buée, Xavier Hanoulle, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U1172 Inserm - U837 (JPArc), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Lille Nord de France (COMUE)-Université de Lille, Hybrigenics [Paris], Hybrigenics, Université de Lille-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U837 (JPArc), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, ANR-16-IDEX-0004,ULNE,ULNE(2016), ANR-18-CE44-0016,ToNIC,Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau(2018), Centre d’Infection et d’Immunité de Lille (CIIL) - U1019 - UMR 8204 (CIIL), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc - U1172 Inserm), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université Lille 2 - Faculté de Médecine, Institut de Chimie des Substances Naturelles (ICSN), Centre National de la Recherche Scientifique (CNRS), Centre de recherche Jean-Pierre Aubert-Neurosciences et Cancer, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Droit et Santé, and Landrieu, Isabelle
Subjects: Genetically modified mouse, Physiology, Cognitive Neuroscience, [SDV]Life Sciences [q-bio], Tau protein, Variable domain of the heavy-chain only antibodies, Mice, Transgenic, tau Proteins, Context (language use), Biochemistry, Epitope, Mice, 03 medical and health sciences, 0302 clinical medicine, Alzheimer Disease, Cell Line, Tumor, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Immunoglobulin Fragments, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, nuclear magnetic resonance spectroscopy, Neurons, 0303 health sciences, biology, Chemistry, tauopathies, Cell Biology, General Medicine, Single-Domain Antibodies, Alzheimer's disease, In vitro, nanobodies, Cell biology, [SDV] Life Sciences [q-bio], Single-domain antibody, Cell culture, biology.protein, Antibody, 030217 neurology & neurosurgery
Abstract: Tau is a neuronal protein linked to pathologies called tauopathies, including Alzheimer's disease. In Alzheimer's disease, tau aggregates into filaments, leading to the observation of intraneuronal fibrillary tangles. Molecular mechanisms resulting in tau aggregation and in tau pathology spreading through the brain regions are still not fully understood. New tools are thus needed to decipher tau pathways involved in the diseases. In this context, a family of novel single domain antibody fragments, or VHHs, directed against tau were generated and characterized. Among the selected VHHs obtained from screening of a synthetic library, a family of six VHHs shared the same CDR3 recognition loop and recognized the same epitope, located in the C-terminal domain of tau. Affinity parameters characterizing the tau/VHHs interaction were next evaluated using surface plasmon resonance spectroscopy. The equilibrium constants KD were in the micromolar range, but despite conservation of the CDR3 loop sequence, a range of affinities was observed for this VHH family. One of these VHHs, named F8-2, was additionally shown to bind tau upon expression in a neuronal cell line model. Optimization of VHH F8-2 by yeast two-hybrid allowed the generation of an optimized VHH family characterized by lower KD than that of the F8-2 wild-type counterpart, and recognizing the same epitope. The optimized VHHs can also be used as antibodies for detecting tau in transgenic mice brain tissues. These results validate the use of these VHHs for in vitro studies, but also their potential for in-cell expression and assays in mouse models, to explore the mechanisms underlying tau physiopathology.
Published: 2019
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30. The elusive tau molecular structures: can we translate the recent breakthroughs into new targets for intervention?

Author: Isabelle Landrieu, Yann Fichou, Joke Verelst, Eugeen Vanmechelen, Youssra K. Al-Hilaly, Luc Buée, Nick Geukens, François Devred, Audrey Perrotin, Miguel Medina, Louise C. Serpell, Caroline Smet-Nocca, Philipp O. Tsvetkov, Bernard Hanseeuw, Joris Winderickx, Department of Chemistry and Biochemistry [Los Angeles], University of California [Los Angeles] (UCLA), University of California (UC)-University of California (UC), University of Sussex, Institut de neurophysiopathologie (INP), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Centre de Recherches en Oncologie biologique et Oncopharmacologie (CRO2), Aix Marseille Université (AMU)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Institut National de la Santé et de la Recherche Médicale (INSERM), Catholic University of Leuven - Katholieke Universiteit Leuven (KU Leuven), ADx Neurosciences NV, Neuropsychologie cognitive et neuroanatomie fonctionnelles de la mémoire humaine, Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-École Pratique des Hautes Études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM), Cliniques Universitaires Saint-Luc [Bruxelles], Centro de Investigacion Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Instituto de Salud Carlos III [Madrid] (ISC), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U837 (JPArc), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, University of California-University of California, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), École pratique des hautes études (EPHE)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Center for Biomedical Research in Neurodegenerative Diseases Network [Madrid, Spain] (CIBERNED ), Queen Sofia Foundation Alzheimer Center [Madrid, Spain] (CIEN Foundation), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc - U837 Inserm), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université Lille 2 - Faculté de Médecine, UCL - SSS/IONS/NEUR - Clinical Neuroscience, UCL - (SLuc) Service de neurologie, Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Aix Marseille Université (AMU), Normandie Université (NU)-Normandie Université (NU)-École pratique des hautes études (EPHE), Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U1172 Inserm - U837 (JPArc), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Lille Nord de France (COMUE)-Université de Lille
Subjects: 0301 basic medicine, Computer science, [SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology, Review, lcsh:RC346-429, Translational Research, Biomedical, 0302 clinical medicine, PHOSPHORYLATED TAU, Alzheimer's disease diagnosis, CORE, Alzheimer’s disease, tau structure, tau aggregation, biology, PAIRED HELICAL FILAMENTS, Brain, Alzheimer's disease, 3. Good health, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, Tauopathies, Tau phosphorylation, Life Sciences & Biomedicine, PROTEIN-TAU, Amyloid, NEUROFIBRILLARY PATHOLOGY, Tau protein, tau Proteins, [SDV.CAN]Life Sciences [q-bio]/Cancer, Pathology and Forensic Medicine, 03 medical and health sciences, Cellular and Molecular Neuroscience, Protein Aggregates, CEREBROSPINAL-FLUID, Tau aggregation, mental disorders, Alzheimer’s disease diagnosis, Animals, Humans, lcsh:Neurology. Diseases of the nervous system, Science & Technology, Neurosciences, IN-VITRO, AGGREGATION, Tau structure, EMISSION-TOMOGRAPHY TRACER, 030104 developmental biology, biology.protein, Paired helical filaments, ALZHEIMERS-DISEASE BRAIN, Neurology (clinical), Neurosciences & Neurology, Neuroscience, 030217 neurology & neurosurgery
Abstract: Insights into tau molecular structures have advanced significantly in recent years. This field has been the subject of recent breakthroughs, including the first cryo-electron microscopy structures of tau filaments from Alzheimer's and Pick's disease inclusions, as well as the structure of the repeat regions of tau bound to microtubules. Tau structure covers various species as the tau protein itself takes many forms. We will here address a range of studies that help to define the many facets of tau protein structures and how they translate into pathogenic forms. New results shed light on previous data that need now to be revisited in order to up-date our knowledge of tau molecular structure. Finally, we explore how these data can contribute the important medical aspects of this research - diagnosis and therapeutics. ispartof: ACTA NEUROPATHOLOGICA COMMUNICATIONS vol:7 issue:1 ispartof: location:England status: published
Published: 2019
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31. Nuclear Magnetic Resonance Spectroscopy Insights into Tau Structure in Solution: Impact of Post-translational Modifications

Author: Alessia Lasorsa, Xavier Hanoulle, Elian Dupre, Caroline Smet-Nocca, Clément Danis, João Filipe Neves, Isabelle Landrieu, Robert Schneider, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: Tau protein, tau Proteins, Isomerase, Protein Structure, Secondary, 03 medical and health sciences, 0302 clinical medicine, Humans, Peptide bond, [CHIM]Chemical Sciences, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, 030212 general & internal medicine, Phosphorylation, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Spectroscopy, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, Quantitative Biology::Biomolecules, biology, Chemistry, Trans conformation, Nuclear magnetic resonance spectroscopy, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Tau phosphorylation, Posttranslational modification, Biophysics, biology.protein, Protein Processing, Post-Translational
Abstract: Although Tau is an intrinsically disordered protein, some level of structure can still be defined, corresponding to short stretches of dynamic secondary structures and a preferential global fold described as an ensemble of conformations. These structures can be modified by Tau phosphorylation, and potentially other post-translational modifications. The analytical capacity of Nuclear Magnetic Resonance (NMR) spectroscopy provides the advantage of offering a residue-specific view of these modifications, allowing to link specific sites to a particular structure. The cis or trans conformation of X-Proline peptide bonds is an additional characteristic parameter of Tau structure that is targeted and modified by prolyl cis/trans isomerases. The challenge in molecular characterization of Tau lies in being able to link structural parameters to functional consequences in normal functions and dysfunctions of Tau, including potential misfolding on the path to aggregation and/or perturbation of the interactions of Tau with its many molecular partners. Phosphorylation of Ser and Thr residues has the potential to impact the local and global structure of Tau.
Published: 2019
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32. Prévention des mécanismes de nucléation et propagation de tau par immunothérapie anti-tau ciblant un épitope central

Author: Raphaëlle Caillierez, Mathieu Schmitt, Clément Danis, Martin Citron, Sébastien Carrier, Marie Albert, Luc Buée, Anne Michel, Isabelle Landrieu, Patrick Downey, Georges Mairet-Coello, Sarah Lieger, Emilie Skrobala, Morvane Colin, Jean-Philippe Courade, BUEE, Luc, Appel à projets générique - Transfert différentiel intercellulaire des assemblages de Tau - - SPREADTAU2014 - ANR-14-CE13-0031 - Appel à projets générique - VALID, APPEL À PROJETS GÉNÉRIQUE 2018 - Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau - - ToNIC2018 - ANR-18-CE44-0016 - AAPG2018 - VALID, Alzheimer & Tauopathies [JPArc], Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer - U837 (JPArc), Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Université Lille Nord de France (COMUE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, UCB BioPharma [Braine l’Alleud, Belgium], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Centre Mémoire de Ressources et de Recherche -CMRR [Lille-Bailleul], Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Hôpital Roger Salengro, Evaluation des technologies de santé et des pratiques médicales - ULR 2694 (METRICS), Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), This work was supported by grants from UCB Biopharma SPRL, the program Investissement d’avenir LabEx (laboratory excellence) DISTALZ (Development of Innovative Strategies for a Transdisciplinary approach to ALZheimer’s disease), SPREADTAU ANR-14-CE13–0031, TONIC ANR-18-CE44–0016 and SUSTAIN TUNABLE (I-SITE ULNE). Our laboratories are also supported by LiCEND (Lille Centre of Excellence in Neurodegenerative Disorders), CNRS, Inserm, Me´tropole Europe´enne de Lille, Univ. Lille and DN2M., ANR-14-CE13-0031,SPREADTAU,Transfert différentiel intercellulaire des assemblages de Tau(2014), ANR-18-CE44-0016,ToNIC,Caractérisation moléculaire et cellulaire de nanobodies dirigés contre Tau(2018), CHU Lille, CNRS, Inserm, Université de Lille, Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc) - U1172, Unité de Glycobiologie Structurale et Fonctionnelle (UGSF) - UMR 8576, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], and Lille Neurosciences & Cognition (LilNCog) - U 1172
Subjects: Male, 0301 basic medicine, Genetically modified mouse, medicine.medical_treatment, Tau protein, Cell, Mice, Transgenic, tau Proteins, Antibodies, Epitope, Epitopes, 03 medical and health sciences, 0302 clinical medicine, Alzheimer Disease, In vivo, mental disorders, propagation, medicine, Animals, Immunologic Factors, [SDV.NEU] Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], biology, Propagation, Seeding, Alzheimer\u2019s disease, Immunotherapy, Brain, Neurofibrillary Tangles, Original Articles, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, In vitro, 3. Good health, Disease Models, Animal, 030104 developmental biology, medicine.anatomical_structure, Tauopathies, Disease Progression, biology.protein, Cancer research, Female, [SDV.NEU]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC], Neurology (clinical), immunotherapy, Antibody, [SDV.IMM.IMM] Life Sciences [q-bio]/Immunology/Immunotherapy, Alzheimer’s disease, 030217 neurology & neurosurgery, seeding
Abstract: Immunotargeting of extracellular tau could slow the prion-like spreading of neurodegeneration. Albert et al. report that antibodies that recognize a central epitope on tau are the most effective at blocking both seeding and propagation of tau pathological species in transgenic mouse models seeded by materials derived from Alzheimer’s disease brains., Tauopathies are neurodegenerative diseases characterized by the intraneuronal accumulation of aggregated tau. The staging of this neurodegenerative process is well established for Alzheimer’s disease as well as for other tauopathies. The stereotypical pattern of tau pathology in these diseases is consistent with the hypothesis that the tau protein can spread in a ‘prion-like’ manner. It proposes that extracellular pathological tau species can transmit pathology from cell to cell. Accordingly, by targeting these spreading species with therapeutic antibodies one should be able to slow or halt the progression of tau pathology. To be effective, antibodies should neutralize the pathological species present in Alzheimer’s disease brains and block their cell-to-cell spread. To evaluate both aspects, tau antibody D, which recognizes an epitope in the central region of tau, and was selected for its outstanding ability to block tau seeding in cell based assays, was used in this study. Here, we addressed two fundamental questions: (i) can this anti-tau antibody neutralize the pathological species present in Alzheimer’s disease brains; and (ii) can it block the cell-to-cell spread of tau seeds in vivo? First, antibody D effectively prevented the induction of tau pathology in the brains of transgenic mice that had been injected with human Alzheimer’s disease brain extracts, showing that it could effectively neutralize the pathological species present in these extracts. Second, by using K18 P301L tau fibrils to induce pathology, we further demonstrated that antibody D was also capable of blocking the progression of tau pathology to distal brain regions. In contrast, an amino-terminal tau antibody, which was less effective at blocking tau seeding in vitro showed less efficacy in reducing Alzheimer’s disease patient tau driven pathology in the transgenic mouse model. We did not address whether the same is true for a spectrum of other amino-terminal antibodies that were tested in vitro. These data highlight important differences between tau antibodies and, when taken together with other recently published data, suggest that epitope may be important for function.
Published: 2019
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33. BIN1 recovers tauopathy-induced long-term memory deficits in mice and interacts with Tau through Thr348 phosphorylation

Author: Mikko Hiltunen, Idir Malki, Alessia Lasorsa, Amandine Flaig, Maxime Sartori, Yann Herault, Shruti Desai, Isabelle Landrieu, Mikael Marttinen, Jean-Charles Lambert, Tiago Mendes, Julien Chapuis, Philippe Amouyel, Jocelyn Laporte, Devrim Kilinc, Benoit Deprez, Anais-Camille Vreulx, Nicolas Malmanche, Laurent Pradier, Petra Mäkinen, Florence Leroux, Adrien Herldean, Damien Marechal, and François-Xavier Cantrelle
Subjects: Genetically modified mouse, 0303 health sciences, Long-term memory, Kinase, Hippocampus, Biology, medicine.disease, 3. Good health, Cell biology, Calcineurin, 03 medical and health sciences, 0302 clinical medicine, medicine, Phosphorylation, Tauopathy, Alzheimer's disease, 030217 neurology & neurosurgery, 030304 developmental biology
Abstract: The bridging integrator 1 gene (BIN1) is a major genetic risk factor for Alzheimer’s disease (AD). In this report, we investigated how BIN1-dependent pathophysiological processes might be associated with Tau. We first generated a cohort of control and transgenic mice either overexpressing human MAPT (TgMAPT) or both human MAPT and BIN1 (TgMAPT;TgBIN1), which we followed-up from 3 to 15 months. In TgMAPT;TgBIN1 mice short-term memory deficits appeared earlier than in TgMAPT mice; however – unlike TgMAPT mice – TgMAPT;TgBIN1 mice did not exhibit any long-term or spatial memory deficits for at least 15 months. After sacrifice of the cohort at 18 months, immunohistochemistry revealed that BIN1 overexpression prevents both Tau mislocalization and somatic inclusion in the hippocampus, where an increase in BIN1-Tau interaction was also observed. We then sought mechanisms controlling the BIN1-Tau interaction. We developed a high-content screening approach to characterize modulators of the BIN1-Tau interaction in an agnostic way (1,126 compounds targeting multiple pathways), and we identified – among others – an inhibitor of Calcineurin, a Ser/Thr phosphatase. We determined that Calcineurin dephosphorylates BIN1 on a Cyclin-dependent kinase phosphorylation site at T348, promoting the open conformation of the neuronal BIN1 isoform. Phosphorylation of this site increases the availability of the BIN1 SH3 domain for Tau interaction, as demonstrated by nuclear magnetic resonance experiments and in primary neurons. Finally, we observed that the levels of the neuronal BIN1 isoform were decreased in AD brains, whereas phospho-BIN1(T348):BIN1 ratio was increased, suggesting a compensatory mechanism. In conclusion, our data support the idea that BIN1 modulates the AD risk through an intricate regulation of its interaction with Tau. Alteration in BIN1 expression or activity may disrupt this regulatory balance with Tau and have direct effects on learning and memory.
Published: 2018
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34. Direct Crosstalk Between

Author: Gwendoline, Bourré, François-Xavier, Cantrelle, Amina, Kamah, Béatrice, Chambraud, Isabelle, Landrieu, and Caroline, Smet-Nocca
Subjects: Endocrinology, O-GlcNAcylation, NMR spectroscopy, phosphorylation, mental disorders, Tau protein, macromolecular substances, Original Research, crosstalk
Abstract: The formation of intraneuronal fibrillar inclusions of tau protein is associated with several neurodegenerative diseases referred to as tauopathies including Alzheimer's disease (AD). A common feature of these pathologies is hyperphosphorylation of tau, the main component of fibrillar assemblies such as Paired Helical Filaments (PHFs). O-β-linked N-acetylglucosaminylation (O-GlcNAcylation) is another important posttranslational modification involved in regulation of tau pathophysiology. Among the benefits of O-GlcNAcylation, modulation of tau phosphorylation levels and inhibition of tau aggregation properties have been described while decreased O-GlcNAcylation could be involved in the raise of tau phosphorylation associated with AD. However, the molecular mechanisms at the basis of these observations remain to be defined. In this study, we identify by NMR spectroscopy O-GlcNAc sites in the longest isoform of tau and investigate the direct role of O-GlcNAcylation on tau phosphorylation and conversely, the role of phosphorylation on tau O-GlcNAcylation. We show here by a systematic examination of the quantitative modification patterns by NMR spectroscopy that O-GlcNAcylation does not modify phosphorylation of tau by the kinase activity of ERK2 or a rat brain extract while phosphorylation slightly increases tau O-GlcNAcylation by OGT. Our data suggest that indirect mechanisms act in the reciprocal regulation of tau phosphorylation and O-GlcNAcylation in vivo involving regulation of the enzymes responsible of phosphate and O-GlcNAc dynamics.
Published: 2018

35. Inhibition of 14-3-3/Tau by Hybrid Small-Molecule Peptides Operating via Two Different Binding Modes

Author: João Neves, Christian Ottmann, Femke A. Meijer, Sebastian A. Andrei, Isabelle Landrieu, Luc Brunsveld, Lech-Gustav Milroy, Technische Universiteit Eindhoven (TU/e), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Eindhoven University of Technology [Eindhoven] (TU/e), Universität Duisburg-Essen [Essen], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Chemical Biology, Université de Lille-Centre National de la Recherche Scientifique (CNRS), and Universität Duisburg-Essen = University of Duisburg-Essen [Essen]
Subjects: 0301 basic medicine, Magnetic Resonance Spectroscopy, Physiology, protein-protein interactions, Peptide, Plasma protein binding, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, SDG 3 – Goede gezondheid en welzijn, Crystallography, X-Ray, Biochemistry, Microtubules, inhibitors, tau, Phosphorylation, tau Proteins/chemistry, 14-3-3, ComputingMilieux_MISCELLANEOUS, media_common, chemistry.chemical_classification, Crystallography, Chemistry, Drug discovery, Neurodegeneration, Neurofibrillary Tangles, General Medicine, Small molecule, 3. Good health, Cell biology, Microtubules/metabolism, ddc:540, Alzheimer Disease/metabolism, Research Article, Protein Binding, Drug, Cognitive Neuroscience, media_common.quotation_subject, Chemie, tau Proteins, Neuroprotection, Protein–protein interaction, drug discovery, 03 medical and health sciences, SDG 3 - Good Health and Well-being, Alzheimer Disease, medicine, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Cell Biology, medicine.disease, 030104 developmental biology, 14-3-3 Proteins, peptide chemistry, X-Ray, protein−protein interactions, 14-3-3 Proteins/chemistry, Neurofibrillary Tangles/metabolism
Abstract: Current molecular hypotheses have not yet delivered marketable treatments for Alzheimer's disease (AD), arguably due to a lack of understanding of AD biology and an overreliance on conventional drug modalities. Protein-protein interactions (PPIs) are emerging drug targets, which show promise for the treatment of, e.g., cancer, but are still underexploited for treating neurodegenerative diseases. 14-3-3 binding to phosphorylated Tau is a promising PPI drug target based on its reported destabilizing effect on microtubules, leading to enhanced neurofibrillary tangle formation as a potential cause of AD-related neurodegeneration. Inhibition of 14-3-3/Tau may therefore be neuroprotective. Previously, we reported the structure-guided development of modified peptide inhibitors of 14-3-3/Tau. Here, we report further efforts to optimize the binding mode and activity of our modified Tau peptides through a combination of chemical synthesis, biochemical assays, and X-ray crystallography. Most notably, we were able to characterize two different high-affinity binding modes, both of which inhibited 14-3-3-binding to full-length PKA-phosphorylated Tau protein in vitro as measured by NMR spectroscopy. Our findings, besides producing useful tool inhibitor compounds for studying 14-3-3/Tau, have enhanced our understanding of the molecular parameters for inhibiting 14-3-3/Tau, which are important milestones toward the establishment of our 14-3-3 PPI hypothesis.
Published: 2018
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36. F2-06-01: MAJOR DIFFERENCES BETWEEN THE SELF-ASSEMBLY, SEEDING BEHAVIOR, AND INTERACTION WITH MODULATORS OF HEPARIN-INDUCED VERSUS IN-VITRO PHOSPHORYLATED TAU

Author: Chunyu Wang, Fuming Zhang, Guy Lippens, Isabelle Huvent, Thomas Schrader, Shiguo Chen, Zizheng Li, Jing Zhao, François-Xavier Cantrelle, Isabelle Landrieu, Robert J. Linhardt, Jianle Chen, Gal Bitan, Jing Di, Caroline Smet-Nocca, Béatrice Chambraud, Frank-Gerrit Klärner, and Clément Despres
Subjects: Epidemiology, Chemistry, Health Policy, Heparin, In vitro, Psychiatry and Mental health, Cellular and Molecular Neuroscience, Developmental Neuroscience, Tau phosphorylation, medicine, Biophysics, Seeding, Neurology (clinical), Geriatrics and Gerontology, medicine.drug
Published: 2019
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37. Stabilizer‐Guided Inhibition of Protein–Protein Interactions

Author: Lech‐Gustav Milroy, Maria Bartel, Morkos A. Henen, Seppe Leysen, Joris M. C. Adriaans, Luc Brunsveld, Isabelle Landrieu, and Christian Ottmann
Subjects: General Medicine
Published: 2015
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38. Solution Structure of the N-Terminal Domain of Mediator Subunit MED26 and Molecular Characterization of Its Interaction with EAF1 and TAF7

Author: Zoé Lens, Xavier Hanoulle, Frédérique Dewitte, François-Xavier Cantrelle, Riccardo Peruzzini, Isabelle Landrieu, Marc Aumercier, Elisabeth Ferreira, Jean-Luc Baert, Vincent Villeret, Didier Monté, Alexis Verger, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), The NMR facilities were funded by the Hauts-de-France Regional Council, CNRS, Pasteur Institute of Lille, European Community (FEDER), French Research Ministry and the University of Sciences and Technologies of Lille I. We acknowledge support from the TGE RMN THC (FR-3050, France), FRABio (FR-3688, France), 'Projets Emergents' Nord-Pas-de-Calais Regional Council and Lille NMR and RPE Health and Biology core facility. R.P. was funded by a fellowship from Nord-Pas-de-Calais Regional Council., The FP7 WeNMR (project no. 261572) and H2020 West-Life (project no. 675858) European e-Infrastructure projects are acknowledged for the use of their Web portals., Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), ANR-16-CE12-0021,MEDNET,Le complexe Médiateur: d'un réseau d'interactions protéine-protéine à la régulation génique in vivo(2016), and Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Magnetic Resonance Spectroscopy, Protein Conformation, Protein subunit, RNA polymerase II, Biology, MESH: Transcription Factor TFIID/metabolism, 03 medical and health sciences, chemistry.chemical_compound, NMR spectroscopy, MESH: Protein Conformation, Structural Biology, Transcription (biology), RNA polymerase, Protein Interaction Mapping, Humans, MESH: Mediator Complex/metabolism, MESH: Protein Binding, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, protein structure, Molecular Biology, TATA-Binding Protein Associated Factors, MESH: Transcription Factors/metabolism, MESH: Humans, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], MEDiator complex, MESH: Magnetic Resonance Spectroscopy, MESH: Protein Interaction Mapping, MED26, MESH: Transcription Factor TFIID/chemistry, TAF7, Dissociation constant, Crystallography, MESH: Mediator Complex/chemistry, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030104 developmental biology, protein–protein interaction, chemistry, MESH: TATA-Binding Protein Associated Factors/chemistry, MESH: TATA-Binding Protein Associated Factors/metabolism, Helix, biology.protein, Biophysics, Transcription Factor TFIID, transcription regulation, Protein Binding, Transcription Factors
Abstract: International audience; MED26 is a subunit of Mediator, a large complex central to the regulation of gene transcription by RNA Polymerase II. MED26 plays a role in the switch between the initiation and elongation phases of RNA Polymerase II-mediated transcription process. Regulation of these steps requires successive binding of MED26 N-terminal domain (NTD) to TATA-binding protein-associated factor 7 (TAF7) and Eleven-nineteen lysine-rich in leukemia-Associated Factor 1 (EAF1). In order to investigate the mechanism of regulation by MED26, MED26-NTD structure was solved by NMR, revealing a 4-helix bundle. EAF1 (239-268) and TAF7 (205-235) peptide interactions were both mapped to the same groove formed by H3 and H4 helices of MED26-NTD. Both interactions are characterized by dissociation constants in the 10-μM range. Further experiments revealed a folding-upon-binding mechanism that leads to the formation of EAF1 (N247-S260) and TAF7 (L214-S227) helices. Chemical shift perturbations and nuclear Overhauser enhancement contacts support the involvement of residues I222/F223 in anchoring TAF7 helix to a hydrophobic pocket of MED26-NTD, including residues L48, W80 and I84. In addition, Ala mutations of charged residues located in the C-terminal disordered part of TAF7 and EAF1 peptides affected the binding, with a loss of affinity characterized by a 10-time increase of dissociation constants. A structural model of MED26-NTD/TAF7 complex shows bi-partite components, combining ordered and disordered segments, as well as hydrophobic and electrostatic contributions to the binding. This study provides molecular detail that will help to decipher the mechanistic basis for the initiation to elongation switch-function mediated by MED26-NTD.
Published: 2017
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39. Regulation of the interaction between the neuronal BIN1 isoform 1 and Tau proteins - role of the SH3 domain

Author: Isabelle Landrieu, François-Xavier Cantrelle, Yoann Sottejeau, Guy Lippens, Idir Malki, Jean-Charles Lambert, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP), The NMR facilities were funded by the Nord Region Council, CNRS, Pasteur Institute of Lille, European Community (FEDER), French Research Ministry and Lille University. We acknowledge support from TGE RMN THC (FR-3050, France) and FRABio (Univ. Lille, CNRS, FR 3688). This study was supported by a grant from the LabEx (Laboratory of Excellence), DISTALZ (Development of Innovative Strategies for a Transdisciplinary approach to Alzheimer's disease), by the French government funding agency Agence Nationale de la Recherche ANR-15-CE16-0002 BinAlz and by the Alzheimer's association (BFG-14-318355)., Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), CNRS, Université de Lille, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Facteurs de Risque et Déterminants Moléculaires des Maladies liées au Vieillissement - U 1167 [RID-AGE], and Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 [UGSF]
Subjects: 0301 basic medicine, Models, Molecular, Protein Conformation, alpha-Helical, Alzheimer’s disease, BIN1, nuclearmagnetic resonance spectroscopy, protein–protein interaction, SH3 domain, Tau, MESH: Sequence Homology, Amino Acid, MESH: Adaptor Proteins, Signal Transducing/metabolism, Amino Acid Motifs, MESH: Tumor Suppressor Proteins/genetics, Gene Expression, Peptide, Biochemistry, MESH: Amino Acid Motifs, 0302 clinical medicine, MESH: Adaptor Proteins, Signal Transducing/genetics, MESH: Nuclear Magnetic Resonance, Biomolecular, Protein Isoforms, Cloning, Molecular, MESH: tau Proteins/chemistry, chemistry.chemical_classification, Neurons, biology, MESH: Kinetics, Chemistry, Nuclear Proteins, MESH: Neurons/metabolism, Nuclear magnetic resonance spectroscopy, MESH: Peptides/genetics, MESH: Tumor Suppressor Proteins/chemistry, MESH: tau Proteins/metabolism, Recombinant Proteins, MESH: Tumor Suppressor Proteins/metabolism, MESH: Recombinant Proteins/chemistry, MESH: tau Proteins/genetics, MESH: Peptides/metabolism, Domain (ring theory), MESH: Protein Conformation, beta-Strand, MESH: Protein Isoforms/metabolism, MESH: Models, Molecular, MESH: Nuclear Proteins/genetics, Protein Binding, Gene isoform, MESH: Recombinant Proteins/metabolism, MESH: Gene Expression, MESH: Sequence Alignment, tau Proteins, MESH: Escherichia coli/genetics, Clathrin, Protein–protein interaction, MESH: Peptides/chemistry, 03 medical and health sciences, Escherichia coli, Humans, MESH: Protein Binding, Protein Interaction Domains and Motifs, MESH: Adaptor Proteins, Signal Transducing/chemistry, MESH: Recombinant Proteins/genetics, MESH: Cloning, Molecular, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Nuclear Proteins/metabolism, Molecular Biology, Gene, Nuclear Magnetic Resonance, Biomolecular, Adaptor Proteins, Signal Transducing, MESH: Protein Conformation, alpha-Helical, MESH: Protein Interaction Domains and Motifs, Binding Sites, MESH: Protein Isoforms/genetics, MESH: Humans, Sequence Homology, Amino Acid, Tumor Suppressor Proteins, Cell Biology, MESH: Nuclear Proteins/chemistry, Kinetics, MESH: Neurons/chemistry, 030104 developmental biology, MESH: Binding Sites, MESH: Protein Isoforms/chemistry, Biophysics, biology.protein, MESH: Escherichia coli/metabolism, Protein Conformation, beta-Strand, Peptides, Sequence Alignment, 030217 neurology & neurosurgery
Abstract: International audience; Bridging integrator 1 (bin1) gene is a genetic determinant of Alzheimer's disease (AD) and has been reported to modulate Alzheimer's pathogenesis through pathway(s) involving Tau. The functional impact of Tau/BIN1 interaction as well as the molecular details of this interaction are still not fully resolved. As a consequence, how BIN1 through its interaction with Tau affects AD risk is also still not determined. To progress in this understanding, interaction of Tau with two BIN1 isoforms was investigated using Nuclear Magnetic Resonance spectroscopy. 1H, 15N spectra showed that the C-terminal SH3 domain of BIN1 isoform 1 (BIN1Iso1) is not mobile in solution but locked with the core of the protein. In contrast, the SH3 domain of BIN1 isoform 9 (BIN1Iso9) behaves as an independent mobile domain. This reveals an equilibrium between close and open conformations for the SH3 domain. Interestingly, a 334–376 peptide from the clathrin and AP-2-binding domain (CLAP) domain of BIN1Iso1, which contains a SH3-binding site, is able to compete with BIN1-SH3 intramolecular interaction. For both BIN1 isoforms, the SH3 domain can interact with Tau(210–240) sequence. Tau(210–240) peptide can indeed displace the intramolecular interaction of the BIN1-SH3 of BIN1Iso1 and form a complex with the released domain. The measured Kd were in agreement with a stronger affinity of Tau peptide. Both CLAP and Tau peptides occupied the same surface on the BIN1-SH3 domain, showing that their interaction is mutually exclusive. These results emphasize an additional level of complexity in the regulation of the interaction between BIN1 and Tau dependent of the BIN1 isoforms.
Published: 2017
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40. Identification of the Tau phosphorylation pattern that drives its aggregation

Author: François-Xavier Cantrelle, Haoling Qi, Yves Jacquot, Guy Lippens, Isabelle Huvent, Isabelle Landrieu, Cillian Byrne, Clément Despres, Béatrice Chambraud, Etienne-Emile Baulieu, Caroline Smet-Nocca, CNRS, Université de Lille, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 [UGSF], Laboratoire des biomolécules [LBM UMR 7203], Université Pierre et Marie Curie - Paris 6 [UPMC], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Petites Molécules de neuroprotection, neurorégénération et remyélinisation, Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés [LISBP], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Université Pierre et Marie Curie - Paris 6 (UPMC), Neuroprotection et neurogénération (UMR788), Neuroprotection et Neurodégénération (UMR788), TGE RMN THC (Tres Grands Equipements Resonance Magnetique Nucleaire Tres Hauts Champs, France) [FR-3050], FRABio (Federation de Recherche 'Biochimie Structurale & Fonctionnelle des Assemblages Biomoleculaires', France [FR 3688], MetaToul [MetaboHUB-AR-11-INBS-0010], Université de Lille-Centre National de la Recherche Scientifique (CNRS), Landrieu, Isabelle, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), and Lippens, Guy
Subjects: 0301 basic medicine, Models, Molecular, Threonine, inorganic chemicals, Phosphorylation sites, Magnetic Resonance Spectroscopy, Tau, phosphorylation, aggregation, Alzheimer's disease, NMR, tau Proteins, Biotechnologies, macromolecular substances, Protein Aggregation, Pathological, environment and public health, Rats, Sprague-Dawley, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, Microscopy, Electron, Transmission, Protein Domains, Alzheimer’s disease, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Serine, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, ComputingMilieux_MISCELLANEOUS, Multidisciplinary, Binding Sites, Kinase, Nuclear magnetic resonance spectroscopy, Biological Sciences, Fluorescence, In vitro, Peptide Fragments, 3. Good health, enzymes and coenzymes (carbohydrates), 030104 developmental biology, chemistry, Biochemistry, Tau phosphorylation, Phosphorylation, bacteria, Thioflavin, 030217 neurology & neurosurgery
Abstract: International audience; Determining the functional relationship between Tau phosphorylation and aggregation has proven a challenge owing to the multiple potential phosphorylation sites and their clustering in the Tau sequence. We use here in vitro kinase assays combined with NMR spectroscopy as an analytical tool to generate well-characterized phosphorylated Tau samples and show that the combined phosphorylation at the Ser202/Thr205/Ser208 sites, together with absence of phosphorylation at the Ser262 site, yields a Tau sample that readily forms fibers, as observed by thioflavin T fluorescence and electron microscopy. On the basis of conformational analysis of synthetic phosphorylated peptides, we show that aggregation of the samples correlates with destabilization of the turn-like structure defined by phosphorylation of Ser202/Thr205.
Published: 2017
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41. The Study of Posttranslational Modifications of Tau Protein by Nuclear Magnetic Resonance Spectroscopy: Phosphorylation of Tau Protein by ERK2 Recombinant Kinase and Rat Brain Extract, and Acetylation by Recombinant Creb-Binding Protein

Author: Jeremy Gunawardena, Caroline Smet-Nocca, Isabelle Landrieu, François-Xavier Cantrelle, Haoling Qi, Béatrice Chambraud, Clément Despres, Guy Lippens, Sudhakaran Prabakaran, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Harvard Medical School [Boston] (HMS), Petites Molécules de neuroprotection, neurorégénération et remyélinisation, Université Paris-Sud - Paris 11 (UP11)-Institut National de la Santé et de la Recherche Médicale (INSERM), Nocca Smet, Caroline, ANR-11-LABX-0009,DISTALZ,Développement de stratégies innovantes pour une approche transdisciplinaire de la maladie d'Alzheime(2011), and Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Recombinant proteins, biology, Kinase, Chemistry, Tau protein, Creb-binding protein, Acetylation, Nuclear magnetic resonance spectroscopy, ERK kinase, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, 03 medical and health sciences, 030104 developmental biology, 0302 clinical medicine, NMR spectroscopy, Biochemistry, Acetyltransferase, biology.protein, Phosphorylation, Kinase activity, CREB-binding protein, 030217 neurology & neurosurgery
Abstract: Nuclear magnetic resonance (NMR) spectroscopy can be used as an analytical tool to investigate posttranslational modifications of protein. NMR is a valuable tool to map the interaction regions of protein partners. Here, we present protocols that have been developed in the course of our studies of the neuronal Tau protein. Tau is found aggregated in the neurons of Alzheimer’s disease patients. Development of the disease is accompanied by increased, abnormal phosphorylation and acetylation of Tau. We have used NMR to investigate how these posttranslational modifications of Tau affect the interactions with its partners. We present here detailed protocols of in vitro phosphorylation of Tau by recombinant kinase, ERK2, or kinase activity of rat brain extracts, and acetylation by recombinant Creb-binding protein (CBP) acetyltransferase. The analytical characterization of the modified Tau by NMR spectroscopy is additionally described.
Published: 2017
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42. NMR reveals the intrinsically disordered domain 2 of NS5A protein as an allosteric regulator of the hepatitis C virus RNA polymerase NS5B

Author: Isabelle Landrieu, Robert Schneider, Guy Lippens, Hélène Launay, Xavier Hanoulle, Luiza M. Bessa, François-Xavier Cantrelle, Marie Dujardin, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), CNRS (IR RMN THC) [FR-3050], ANR-11-JSV8-005, CNRS, Université de Lille, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 [UGSF], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Université de Lille-Centre National de la Recherche Scientifique (CNRS), Schneider, Robert, and Hanoulle, Xavier
Subjects: Models, Molecular, 0301 basic medicine, RNA-protein interaction, Protein Conformation, viruses, Hepacivirus, Viral Nonstructural Proteins, Biochemistry, Protein Refolding, protein-protein interaction, chemistry.chemical_compound, RNA-Protein Interaction, RNA polymerase, chimiothérapie, Enzyme Inhibitors, Polymerase, ComputingMilieux_MISCELLANEOUS, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], virus diseases, Molecular Bases of Disease, hépatite c, Recombinant Proteins, 3. Good health, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Allosteric Site, nuclear magnetic resonance (NMR), Allosteric regulation, Médecine humaine et pathologie, Antiviral Agents, arn, 03 medical and health sciences, Point Mutation, Protein Interaction Domains and Motifs, Isoleucine, NS5A, Nuclear Magnetic Resonance, Biomolecular, Molecular Biology, NS5B, Oligoribonucleotides, génome, RNA, Cell Biology, Triazoles, biochemical phenomena, metabolism, and nutrition, RNA-Dependent RNA Polymerase, Peptide Fragments, digestive system diseases, allosteric regulation, Intrinsically Disordered Proteins, 030104 developmental biology, Solubility, Allosteric enzyme, chemistry, Pyrones, protein dynamic, Mutagenesis, Site-Directed, biology.protein, Biophysics, Human health and pathology, Gene Deletion
Abstract: International audience; Non-structural protein 5B (NS5B) is the RNA-dependent RNA polymerase that catalyzes replication of the hepatitis C virus (HCV) RNA genome and therefore is central for its life cycle. NS5B interacts with the intrinsically disordered domain 2 of NS5A (NS5A-D2), another essential multifunctional HCV protein that is required for RNA replication. As a result, these two proteins represent important targets for anti-HCV chemotherapies. Despite this importance and the existence of NS5B crystal structures, our understanding of the conformational and dynamic behavior of NS5B in solution and its relationship with NS5A-D2 remains incomplete. To address these points, we report the first detailed NMR spectroscopic study of HCV NS5B lacking its membrane anchor (NS5B(21)). Analysis of constructs with selective isotope labeling of the 1 methyl groups of isoleucine side chains demonstrates that, in solution, NS5B(21) is highly dynamic but predominantly adopts a closed conformation. The addition of NS5A-D2 leads to spectral changes indicative of binding to both allosteric thumb sites I and II of NS5B(21) and induces long-range perturbations that affect the RNA-binding properties of the polymerase. We compared these modifications with the short- and long-range effects triggered in NS5B(21) upon binding of filibuvir, an allosteric inhibitor. We demonstrate that filibuvir-bound NS5B(21) is strongly impaired in the binding of both NS5A-D2 and RNA. NS5A-D2 induces conformational and functional perturbations in NS5B similar to those triggered by filibuvir. Thus, our work highlights NS5A-D2 as an allosteric regulator of the HCV polymerase and provides new insight into the dynamics of NS5B in solution.
Published: 2017
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43. A functional fragment of Tau forms fibers without the need for an intermolecular cysteine bridge

Author: Amina Kamah, Isabelle Landrieu, Isabelle Huvent, Guy Lippens, Christian Slomianny, François-Xavier Cantrelle, Nicolas Barois, Caroline Smet-Nocca, Université Lille Nord de France (COMUE), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Plateforme BioImaging Center Lille (BICeL), Plateformes Lilloises en Biologie et Santé - UMS 2014 - US 41 (PLBS), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Physiologie Cellulaire : Canaux ioniques, inflammation et cancer - U 1003 (PHYCELL), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université de Lille, Plate-forme BICeL (IFR142), Physiologie Cellulaire (PHYCEL) - U1003 (PHYCEL), Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Lille Nord de France (COMUE), Université de Lille-Centre National de la Recherche Scientifique (CNRS), and Plateformes Lilloises en Biologie et Santé - UAR 2014 - US 41 (PLBS)
Subjects: Magnetic Resonance Spectroscopy, Biophysics, tau Proteins, macromolecular substances, Fibril, Biochemistry, law.invention, Aggregation, 03 medical and health sciences, chemistry.chemical_compound, NMR spectroscopy, 0302 clinical medicine, law, Microtubule, mental disorders, Electron microscopy, Humans, Cysteine, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Binding site, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, 0303 health sciences, Heparin, Cell Biology, Nuclear magnetic resonance spectroscopy, Fluorescence, Recombinant Proteins, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Crystallography, nervous system, chemistry, Nucleation, Thioflavin, Tau, Electron microscope, 030217 neurology & neurosurgery
Abstract: International audience; We study the aggregation of a fragment of the neuronal protein Tau that contains part of the proline rich domain and of the microtubule binding repeats. When incubated at 37 °C with heparin, the fragment readily forms fibers as witnessed by Thioflavin T fluorescence. Electron microscopy and NMR spectroscopy show bundled ribbon like structures with most residues rigidly incorporated in the fibril. Without its cysteines, this fragment still forms fibers of a similar morphology, but with lesser Thioflavin T binding sites and more mobility for the C-terminal residues.
Published: 2014
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44. Nanobodies (VHHs) for targeting tau in Alzheimer's disease and tauopathies

Author: Morvane Colin, François-Xavier Cantrelle, Xavier Hanoulle, Elian Dupre, Alexis Arrial, Clément Danis, Jean-Christophe Rain, Luc Buée, and Isabelle Landrieu
Subjects: business.industry, General Neuroscience, Medicine, Disease, business, Neuroscience
Published: 2019
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45. The Study of Posttranslational Modifications of Tau Protein by Nuclear Magnetic Resonance Spectroscopy: Phosphorylation of Tau Protein by ERK2 Recombinant Kinase and Rat Brain Extract, and Acetylation by Recombinant Creb-Binding Protein

Author: Haoling, Qi, Clément, Despres, Sudhakaran, Prabakaran, François-Xavier, Cantrelle, Béatrice, Chambraud, Jeremy, Gunawardena, Guy, Lippens, Caroline, Smet-Nocca, and Isabelle, Landrieu
Subjects: Mitogen-Activated Protein Kinase 1, Neurons, Alzheimer Disease, Animals, Acetylation, tau Proteins, Phosphorylation, CREB-Binding Protein, Nuclear Magnetic Resonance, Biomolecular, Protein Processing, Post-Translational, Recombinant Proteins, Rats
Abstract: Nuclear magnetic resonance (NMR) spectroscopy can be used as an analytical tool to investigate posttranslational modifications of protein. NMR is a valuable tool to map the interaction regions of protein partners. Here, we present protocols that have been developed in the course of our studies of the neuronal Tau protein. Tau is found aggregated in the neurons of Alzheimer's disease patients. Development of the disease is accompanied by increased, abnormal phosphorylation and acetylation of Tau. We have used NMR to investigate how these posttranslational modifications of Tau affect the interactions with its partners. We present here detailed protocols of in vitro phosphorylation of Tau by recombinant kinase, ERK2, or kinase activity of rat brain extracts, and acetylation by recombinant Creb-binding protein (CBP) acetyltransferase. The analytical characterization of the modified Tau by NMR spectroscopy is additionally described.
Published: 2016

46. A β-Turn Motif in the Steroid Hormone Receptor’s Ligand-Binding Domains Interacts with the Peptidyl-prolyl Isomerase (PPIase) Catalytic Site of the Immunophilin FKBP52

Author: François-Xavier Cantrelle, Guy Lippens, Morkos A. Henen, Haoling Qi, Julien Giustiniani, Amina Kamah, Yves Jacquot, Isabelle Landrieu, Mathilde Belnou, Etienne-Emile Baulieu, Cillian Byrne, Béatrice Chambraud, Université Pierre et Marie Curie - Paris 6 (UPMC), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Neuroprotection et neurogénération (UMR788), Neuroprotection et Neurodégénération (UMR788), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Region Nord, Institut Pasteur of Lille, European Union (FEDER), French Research Ministry, Universite des Sciences et Technologies de Lille 1, University Pierre & Marie Curie (Paris 6) [ED406], Centre National de la Recherche Scientifique (CNRS), Ecole Normale Superieure (ENS), Institut Baulieu, Alanna Schepartz, European Project: 286418,EC:FP7:PEOPLE,FP7-PEOPLE-2011-IAPP,14-3-3STABS(2012), and Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Steroid hormone receptor, Stereochemistry, medicine.medical_treatment, Receptors, Cytoplasmic and Nuclear, Isomerase, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, Biology, Ligands, 010402 general chemistry, 01 natural sciences, Biochemistry, 03 medical and health sciences, Catalytic Domain, Prolyl isomerase, medicine, Binding site, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Receptor, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, Binding Sites, Peptidylprolyl Isomerase, FKBP52, 0104 chemical sciences, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Steroid hormone, 030104 developmental biology, Thermodynamics, Steroids, FKBP1A, Protein Binding
Abstract: International audience; The immunophilin FKBP52 interacts with nuclear steroid hormone receptors. Studying the crystal structure of human estrogen receptor alpha (hER alpha) and using nuclear magnetic resonance, we show here that the short V(364)PGF(367) sequence, which is located within its ligand-binding domain and adopts a type II beta-turn conformation in the protein, binds the peptidyl-prolyl isomerase (PPIase or rotamase) FK1 domain of FKBP52. Interestingly, this turn motif displays strong similarities with the FKBP52 FK1 domain-binding moiety of macrolide immunomodulators such as rapamycin and GPI-1046, an immunophilin ligand with neuroprotective characteristics. An increase in the hydrophobicity of the residue preceding the proline and cyclization of the VPGF peptide strengthen its recognition by the FK1 domain of FKBP52. Replacement of the Pro residue with a dimethylproline also enhances this interaction. Our study not only contributes to a better understanding of how the interaction between the FK1 domain of FKBP52 and steroid hormone receptors most likely works but also opens new avenues for the synthesis of FKBP52 FK1 peptide ligands appropriate for the control of hormone-dependent physiological mechanisms or of the functioning of the Tau protein. Indeed, it has been shown that FKBP52 is involved in the intraneuronal dynamics of the Tau protein.
Published: 2016
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47. NMR Meets Tau: Insights into Its Function and Pathology

Author: Clément Despres, Isabelle Landrieu, Haoling Qi, Isabelle Huvent, Guy Lippens, Neha S. Gandhi, Benoît Gigant, Caroline Smet, Juan José Marín López, Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), School of Mathematical Sciences, Queensland University of Technology, School of Mathematical Sciences [Brisbane], Queensland University of Technology [Brisbane] (QUT)-Queensland University of Technology [Brisbane] (QUT), Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés ( LISBP ), Institut National de la Recherche Agronomique ( INRA ) -Institut National des Sciences Appliquées - Toulouse ( INSA Toulouse ), Institut National des Sciences Appliquées ( INSA ) -Institut National des Sciences Appliquées ( INSA ) -Centre National de la Recherche Scientifique ( CNRS ), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 ( UGSF ), Institut National de la Recherche Agronomique ( INRA ) -Université de Lille-Centre National de la Recherche Scientifique ( CNRS ), School of Mathematical Sciences, Queensland University of Technology [Brisbane] ( QUT ), Biochimie Structurale des Microtubules, des Kinésines et de leurs Cargos ( MIKICA ), Département Biochimie, Biophysique et Biologie Structurale ( B3S ), Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Institut de Biologie Intégrative de la Cellule ( I2BC ), Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ) -Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ), Sciences Department-Chemistry, Pontificia Universidad Católica del Perú ( PUCP ), Queensland University of Technology [Brisbane] (QUT), Biochimie Structurale des Microtubules, des Kinésines et de leurs Cargos (MIKICA), Département Biochimie, Biophysique et Biologie Structurale (B3S), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay-Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Saclay, Pontificia Universidad Católica del Perú (PUCP), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Université Paris-Sud - Paris 11 (UP11)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Queensland University of Technology [Brisbane] ( QUT ) -Queensland University of Technology [Brisbane] ( QUT ), Université Paris-Saclay-Centre National de la Recherche Scientifique ( CNRS ) -Commissariat à l'énergie atomique et aux énergies alternatives ( CEA ) -Université Paris-Sud - Paris 11 ( UP11 ), CNRS, Université de Lille, Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés [LISBP], Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 [UGSF], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Queensland University of Technology [Brisbane] [QUT], Institut de Biologie Intégrative de la Cellule [I2BC], Pontificia Universidad Católica del Perú = Pontifical Catholic University of Peru [PUCP], and Lippens, Guy
Subjects: 0301 basic medicine, [SDV]Life Sciences [q-bio], Tau protein, lcsh:QR1-502, tau Proteins, Review, Intrinsically disordered proteins, Biochemistry, protein interactions, lcsh:Microbiology, Protein–protein interaction, 03 medical and health sciences, [ SDV.BBM.BC ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 0302 clinical medicine, NMR spectroscopy, mental disorders, Tau, intrinsically disordered protein, tubulin, aggregation, phosphorylation, protein/protein interactions, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Molecular Biology, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, biology, [ SDV ] Life Sciences [q-bio], Chemistry, Kinase, protein, Nuclear magnetic resonance spectroscopy, 3. Good health, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Intrinsically Disordered Proteins, 030104 developmental biology, Tubulin, biology.protein, Biophysics, Phosphorylation, Protein Processing, Post-Translational, 030217 neurology & neurosurgery, Function (biology)
Abstract: International audience; In this review, we focus on what we have learned from Nuclear Magnetic Resonance (NMR) studies on the neuronal microtubule-associated protein Tau. We consider both the mechanistic details of Tau: the tubulin relationship and its aggregation process. Phosphorylation of Tau is intimately linked to both aspects. NMR spectroscopy has depicted accurate phosphorylation patterns by different kinases, and its non-destructive character has allowed functional assays with the same samples. Finally, we will discuss other post-translational modifications of Tau and its interaction with other cellular factors in relationship to its (dys)function.
Published: 2016
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48. Characterization of Neuronal Tau Protein as a Target of Extracellular Signal-regulated Kinase

Author: Haoling Qi, Jeremy Gunawardena, Guy Lippens, Béatrice Chambraud, François-Xavier Cantrelle, Sudhakaran Prabakaran, Isabelle Landrieu, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Department of Systems Biology, Harvard Medical School [Boston] (HMS), Neuroprotection et neurogénération (UMR788), Neuroprotection et Neurodégénération (UMR788), Laboratoire d'Ingénierie des Systèmes Biologiques et des Procédés (LISBP), Centre National de la Recherche Scientifique (CNRS)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Institut National de la Recherche Agronomique (INRA), CNRS, Université de Lille, Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 [UGSF], Harvard Medical School [Boston] [HMS], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Petites Molécules de neuroprotection, neurorégénération et remyélinisation, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Institut National des Sciences Appliquées - Toulouse (INSA Toulouse), Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Institut National des Sciences Appliquées (INSA)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Landrieu, Isabelle, Institut National des Sciences Appliquées (INSA)-Université Fédérale Toulouse Midi-Pyrénées-Institut National des Sciences Appliquées (INSA)-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Prabakaran, Sudhakaran [0000-0002-6527-1085], and Apollo - University of Cambridge Repository
Subjects: 0301 basic medicine, MAPK/ERK pathway, Amino Acid Motifs, spectroscopie rmn, Biochemistry, environment and public health, protein-protein interaction, neurodégénérescence, Protein phosphorylation, Phosphorylation, Extracellular Signal-Regulated MAP Kinases, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Kinase, Neurodegeneration, Molecular Bases of Disease, 3. Good health, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], maladie d'alzheimer, extracellular signal-regulated kinase (ERK), protéine, cerveau, Tau protein (Tau), Alzheimer's disease, Alzheimer disease, inorganic chemicals, nuclear magnetic resonance (NMR), Tau protein, Médecine humaine et pathologie, Hyperphosphorylation, tau Proteins, Biology, 03 medical and health sciences, interaction Tau protein (Tau), mental disorders, medicine, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, protein phosphorylation, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], protein phosphorylation protein-protein, Nuclear Magnetic Resonance, Biomolecular, Molecular Biology, Cell Biology, medicine.disease, Protein Structure, Tertiary, Rats, enzymes and coenzymes (carbohydrates), 030104 developmental biology, biology.protein, Human health and pathology
Abstract: International audience; Tau neuronal protein has a central role in neurodegeneration and is implicated in Alzheimer disease development. Abnormal phosphorylation of Tau impairs its interaction with other proteins and is associated with its dysregulation in pathological conditions. Molecular mechanisms leading to hyperphosphorylation of Tau in pathological conditions are unknown. Here, we characterize phosphorylation of Tau by extracellular-regulated kinase (ERK2), a mitogen-activated kinase (MAPK) that responds to extracellular signals. Analysis of in vitro phosphorylated Tau by activated recombinant ERK2 with nuclear magnetic resonance spectroscopy (NMR) reveals phosphorylation of 15 Ser/Thr sites. In vitro phosphorylation of Tau using rat brain extract and subsequent NMR analysis identifies the same sites. Phosphorylation with rat brain extract is known to transform Tau into an Alzheimer disease-like state. Our results indicate that phosphorylation of Tau by ERK2 alone is sufficient to produce the same characteristics. We further investigate the mechanism of ERK2 phosphorylation of Tau. Kinases are known to recognize their protein substrates not only by their specificity for a targeted Ser or Thr phosphorylation site but also by binding to linear-peptide motifs called docking sites. We identify two main ERK2 docking sites in Tau sequence using NMR. Our results suggest that ERK2 dysregulation in Alzheimer disease could lead to abnormal phosphorylation of Tau resulting in the pathology of the disease.
Published: 2016
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49. 1H, 15N and 13C assignments of the N-terminal domain of the Mediator complex subunit MED26

Author: Isabelle Landrieu, Frédérique Dewitte, Alexis Verger, Riccardo Peruzzini, Elisabeth Ferreira, Vincent Villeret, François-Xavier Cantrelle, Zoé Lens, Jean-Luc Baert, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), and Université de Lille-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, congenital, hereditary, and neonatal diseases and abnormalities, Stereochemistry, Protein subunit, RNA polymerase II, 010402 general chemistry, 01 natural sciences, Biochemistry, Protein Structure, Secondary, Protein–protein interaction, 03 medical and health sciences, Mediator, Protein Domains, Structural Biology, Transcription (biology), Transcriptional regulation, Amino Acid Sequence, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, Genetics, Mediator Complex, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], MED26, Nuclear magnetic resonance spectroscopy, 0104 chemical sciences, Protein Subunits, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030104 developmental biology, biology.protein
Abstract: MED26 is a subunit of the Mediator, a very large complex involved in regulation of gene transcription by RNA Polymerase II. MED26 regulates the switch between initiation and elongation phases of the transcription. This function requires interaction of its N-terminal domain (NTD) with several protein partners implicated in transcriptional regulation. Molecular details of the structure and interaction mode of MED26 NTD would improve understanding of this complex regulation. As a first step towards structural characterization, sequence specific (1)H, (13)C and (15)N assignments for MED26 NTD was performed based on Nuclear Magnetic Resonance spectroscopy. TALOS+ analysis of the chemical shifts data revealed a domain solely composed of helices. Assignments will be further used to solve NMR structure and dynamics of MED26 NTD and investigate the molecular details of its interaction with protein partners.
Published: 2016
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50. Isomerisation and oligomerization of truncated and mutated tau forms by FKBP52 are independent processes

Author: Julien Giustiniani, Yves Jacquot, Isabelle Landrieu, Béatrice Chambraud, Guy Lippens, François-Xavier Cantrelle, Caroline Smet, Etienne-Emile Baulieu, Isabelle Huvent, Kevin Guillemeau, Amina Kamah, Cillian Byrne, Université Lille Nord de France (COMUE), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut de Recherche Interdisciplinaire [Villeneuve d'Ascq] (IRI), Centre National de la Recherche Scientifique (CNRS)-Université de Lille, Droit et Santé-Université de Lille, Sciences et Technologies, Neuroprotection et neurogénération (UMR788), Neuroprotection et Neurodégénération (UMR788), Peptides, glycoconjugués et métaux en biologie (LBM-E1), Laboratoire des biomolécules (LBM UMR 7203), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Chimie - ENS Paris, École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Département de Chimie - ENS Paris, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Pierre et Marie Curie - Paris 6 (UPMC), ANR (program MALZ-TAF), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université de Lille, Sciences et Technologies-Université de Lille, Droit et Santé-Centre National de la Recherche Scientifique (CNRS), Université Pierre et Marie Curie - Paris 6 (UPMC)-Département de Chimie - ENS Paris, École normale supérieure - Paris (ENS Paris)-École normale supérieure - Paris (ENS Paris)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Département de Chimie - ENS Paris, École normale supérieure - Paris (ENS Paris)-École normale supérieure - Paris (ENS Paris)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Lille-Centre National de la Recherche Scientifique (CNRS), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Département de Chimie - ENS Paris, and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Isomerase activity, Tau protein, Mutant, tau Proteins, Peptide, Protein Aggregation, Pathological, Tacrolimus Binding Proteins, 03 medical and health sciences, 0302 clinical medicine, Isomerism, Structural Biology, mental disorders, Humans, Molecular Biology, FK506 binding, chemistry.chemical_classification, biology, Chemistry, [CHIM.ORGA]Chemical Sciences/Organic chemistry, aggregation, Alzheimer's disease, FKBP52, prolyl cis/trans isomerase activity, 030104 developmental biology, FKBP, Biochemistry, Biophysics, biology.protein, Protein Multimerization, Tau, 030217 neurology & neurosurgery, Heteronuclear single quantum coherence spectroscopy
Abstract: International audience; The aggregation of the neuronal Tau protein is one molecular hallmark of Alzheimer's disease and other related tauopathies, but the precise molecular mechanisms of the aggregation process remain unclear. The FK506 binding protein FKBP52 is able to induce oligomers in the pathogenic Tau P301L mutant and in a truncated form of the wild-type human Tau protein. Here, we investigate whether FKBP52's capacity to induce Tau oligomers depends on its prolyl cis/trans isomerase activity. We find that FKBP52 indeed can isomerize selected prolyl bonds in the different Tau proteins, and that this activity is carried solely by its first FK506 binding domain. Its capacity to oligomerize Tau is however not linked to this PPIase activity. In addition, we identified a novel molecular interaction implying the PHF6 peptide of Tau and the FK1/FK2 domains of FKBP52 independent of FK506 binding; these data point towards a non-catalytic molecular interaction that might govern the effect of FKBP52 on Tau.
Published: 2016
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