112 results on '"Di Cunto, F"'
Search Results
2. A computational approach to regulatory element discovery in eukaryotes
- Author
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Caselle, M., Di Cunto, F., and Provero, P.
- Subjects
Condensed Matter - Disordered Systems and Neural Networks ,Physics - Biological Physics ,Quantitative Biology - Genomics - Abstract
Gene regulation in Eukaryotes is mainly effected through transcription factors binding to rather short recognition motifs generally located upstream of the coding region. We present a novel computational method to identify regulatory elements in the upstream region of Eukaryotic genes. The genes are grouped in sets sharing an overrepresented short motif in their upstream sequence. For each set, the average expression level from a microarray experiment is determined: if this level is significantly higher or lower than the average taken over the whole genome, then the overrepresented motif shared by the genes in the set is likely to play a role in their regulation. We illustrate the method by applying it to the genome of {\it S. cerevisiae}, for which many datasets of microarray experiments are publicly available. Several known binding motifs are correctly recognized by our algorithm, and a new candidate is suggested for experimental verification., Comment: 7 pages, 2 figures
- Published
- 2003
3. Correlating overrepresented upstream motifs to gene expression: a computational approach to regulatory element discovery in eukaryotes
- Author
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Caselle, M., Di Cunto, F., and Provero, P.
- Subjects
Physics - Biological Physics ,Condensed Matter ,Quantitative Biology - Molecular Networks - Abstract
Gene regulation in eukaryotes is mainly effected through transcription factors binding to rather short recognition motifs generally located upstream of the coding region. We present a novel computational method to identify regulatory elements in the upstream region of eukaryotic genes. The genes are grouped in sets sharing an overrepresented short motif in their upstream sequence. For each set, the average expression level from a microarray experiment is determined: If this level is significantly higher or lower than the average taken over the whole genome, then the overerpresented motif shared by the genes in the set is likely to play a role in their regulation. The method was tested by applying it to the genome of Saccharomyces cerevisiae, using the publicly available results of a DNA microarray experiment, in which expression levels for virtually all the genes were measured during the diauxic shift from fermentation to respiration. Several known motifs were correctly identified, and a new candidate regulatory sequence was determined., Comment: Published version available from http://www.biomedcentral.com/1471-2105/3/7
- Published
- 2002
4. Finding regulatory sites from statistical analysis of nucleotide frequencies in the upstream region of eukaryotic genes
- Author
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Caselle, M., Di Cunto, F., Pellegrino, M., and Provero, P.
- Subjects
Physics - Biological Physics ,Condensed Matter ,Quantitative Biology - Genomics - Abstract
We discuss two new approaches to extract relevant biological information on the Transcription Factors (and in particular to identify their binding sequences) from the statistical distribution of oligonucleotides in the upstream region of the genes. Both the methods are based on the notion of a ``regulatory network'' responsible for the various expression patterns of the genes. In particular we concentrate on families of coregulated genes and look for the simultaneous presence in the upstream regions of these genes of the same set of transcription factor binding sites. We discuss two instances which well exemplify the features of the two methods: the coregulation of glycolysis in Drosophila melanogaster and the diauxic shift in Saccharomyces cerevisiae., Comment: 12 pages. Contribution to the proceedings of the International Workshop ``Modelling Bio-medical signals'', Bari, September 2001
- Published
- 2002
5. Analysis of consequences of CIT-K-dependent cytokinesis failure in p53-proficient or p53-deficient mice: Abstract no. 23
- Author
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Bianchi, F. T., Chang, Y. J., Gai, M., Sgro, F., Huttner, W., and Di Cunto, F.
- Published
- 2013
6. Effect of deep space-related conditions on central nervous system performance in multicellular organisms as a function of their genetic background
- Author
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Berto, G, Di Schiavi, E, Cirio, R, Sacchi, R, Giordanengo, S, Monaco, V, and Di Cunto, F
- Published
- 2019
7. Ab initio identification of putative human transcription factor binding sites by comparative genomics
- Author
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Provero P, Di Cunto F, Dieterich C, Herrmann C, Corà D, and Caselle M
- Subjects
Computer applications to medicine. Medical informatics ,R858-859.7 ,Biology (General) ,QH301-705.5 - Abstract
Abstract Background Understanding transcriptional regulation of gene expression is one of the greatest challenges of modern molecular biology. A central role in this mechanism is played by transcription factors, which typically bind to specific, short DNA sequence motifs usually located in the upstream region of the regulated genes. We discuss here a simple and powerful approach for the ab initio identification of these cis-regulatory motifs. The method we present integrates several elements: human-mouse comparison, statistical analysis of genomic sequences and the concept of coregulation. We apply it to a complete scan of the human genome. Results By using the catalogue of conserved upstream sequences collected in the CORG database we construct sets of genes sharing the same overrepresented motif (short DNA sequence) in their upstream regions both in human and in mouse. We perform this construction for all possible motifs from 5 to 8 nucleotides in length and then filter the resulting sets looking for two types of evidence of coregulation: first, we analyze the Gene Ontology annotation of the genes in the set, searching for statistically significant common annotations; second, we analyze the expression profiles of the genes in the set as measured by microarray experiments, searching for evidence of coexpression. The sets which pass one or both filters are conjectured to contain a significant fraction of coregulated genes, and the upstream motifs characterizing the sets are thus good candidates to be the binding sites of the TF's involved in such regulation. In this way we find various known motifs and also some new candidate binding sites. Conclusion We have discussed a new integrated algorithm for the "ab initio" identification of transcription factor binding sites in the human genome. The method is based on three ingredients: comparative genomics, overrepresentation, different types of coregulation. The method is applied to a full-scan of the human genome, giving satisfactory results.
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- 2005
- Full Text
- View/download PDF
8. Comparison of 3 Tfr2-deficient murine models suggests distinct functions for Tfr2-alpha and Tfr2-beta isoforms in different tissues
- Author
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ROETTO A, DI CUNTO F, PELLEGRINO RM, HIRSCH E, AZZOLINO O, BONDI A, DEFILIPPI I, CARTURAN S, MINISCALCO B, RIONDATO F, CILLONI D, SILENGO L, ALTRUDA F, SAGLIO G., CAMASCHELLA , CLARA, Roetto, A, DI CUNTO, F, Pellegrino, Rm, Hirsch, E, Azzolino, O, Bondi, A, Defilippi, I, Carturan, S, Miniscalco, B, Riondato, F, Cilloni, D, Silengo, L, Altruda, F, Camaschella, Clara, and Saglio, G.
- Published
- 2010
9. Tissue-specific control of midbody microtubule stability by Citron kinase through modulation of TUBB3 phosphorylation
- Author
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Sgrò, F, primary, Bianchi, F T, additional, Falcone, M, additional, Pallavicini, G, additional, Gai, M, additional, Chiotto, A M A, additional, Berto, G E, additional, Turco, E, additional, Chang, Y J, additional, Huttner, W B, additional, and Di Cunto, F, additional
- Published
- 2015
- Full Text
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10. Citron-N is a neuronal Rho-associated protein involved in Golgi organization through actin cytoskeleton regulation
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Camera P, da Silva JS, Griffiths G, Giuffrida MG, Ferrara L, Schubert V, Imarisio S, Silengo L, Dotti CG, and Di Cunto F.
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Citron, actin binding proteins, hippocampal neurons ,Rho GTPases ,Golgi ,macromolecular substances ,actin ,Golgi structure - Abstract
The actin cytoskeleton is best known for its role during cellular morphogenesis. However, other evidence suggests that actin is also crucial for the organization and dynamics of membrane organelles such as endosomes and the Golgi complex. As in morphogenesis, the Rho family of small GTPases are key mediators of organelle actin-driven events, although it is unclear how these ubiquitously distributed proteins are activated to regulate actin dynamics in an organelle-specific manner. Here we show that the brain-specific Rho-binding protein Citron-N is enriched at, and associates with, the Golgi apparatus of hippocampal neurons in culture. Suppression of the whole protein or expression of a mutant form lacking the Rho-binding activity results in dispersion of the Golgi apparatus. In contrast, high intracellular levels induce localized accumulation of RhoA and filamentous actin, protecting the Golgi from the rupture normally produced by actin depolymerization. Biochemical and functional analyses indicate that Citron-N controls actin locally by assembling together the Rho effector ROCK-II and the actin-binding, neuron-specific, protein Profilin-IIa (PIIa). Together with recent data on endosomal dynamics, our results highlight the importance of organelle-specific Rho modulators for actin-dependent organelle organization and dynamics.
- Published
- 2003
11. Modulation of Cellular Hsp72 Levels in Undifferentiated and Neuron-Like SH-SY5Y Cells Determines Resistance to Staurosporine-Induced Apoptosis
- Author
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Di Cunto, F, Cheng, L, Smith, DJ, Anderson, RL, Nagley, P, Di Cunto, F, Cheng, L, Smith, DJ, Anderson, RL, and Nagley, P
- Abstract
Increased expression of Hsp72 accompanies differentiation of human neuroblastoma SH-SY5Y cells to neuron-like cells. By modulating cellular levels of Hsp72, we demonstrate here its anti-apoptotic activity both in undifferentiated and neuron-like cells. Thermal preconditioning (43°C for 30 min) induced Hsp72, leading to cellular protection against apoptosis induced by a subsequent treatment with staurosporine. Preconditioned staurosporine-treated cells displayed decreased Bax recruitment to mitochondria and subsequent activation, as well as reduced cytochrome c redistribution from mitochondria. The data are consistent with Hsp72 blocking apoptosis upstream of Bax recruitment to mitochondria. Neuron-like cells (with elevated Hsp72) were more resistant to staurosporine by all measured indices of apoptotic signaling. Use of stable transfectants ectopically expressing moderately elevated levels of Hsp72 revealed that such cells in the undifferentiated state showed enhanced resistance to staurosporine-induced apoptosis, which was even more robust after differentiation to neuron-like cells. Overall, the protective effects of differentiation, thermal preconditioning and ectopic Hsp72 expression were additive. The strong inverse correlation between cellular Hsp72 levels and susceptibility to apoptosis support the notion that Hsp72 acts as a significant neuroprotective factor, enabling post-mitotic neurons to withstand potentially lethal stress that induces apoptosis.
- Published
- 2011
12. Citron rho-interacting kinase, a novel tissue-specific ser/thr kinase encompassing the Rho-Rac-binding protein Citron
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Jimmy Hsiao, Di Cunto F, Enzo Calautti, Emilia Turco, Lydia Ong, Dotto Gp, and Gabrielle Topley
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Keratinocytes ,DNA, Complementary ,Molecular Sequence Data ,Fluorescent Antibody Technique ,Cell Cycle Proteins ,Protein Serine-Threonine Kinases ,Biology ,Mitogen-activated protein kinase kinase ,Biochemistry ,MAP2K7 ,Mice ,Animals ,ASK1 ,Amino Acid Sequence ,Cloning, Molecular ,Kinase activity ,Molecular Biology ,Base Sequence ,Sequence Homology, Amino Acid ,MAP kinase kinase kinase ,Cyclin-dependent kinase 4 ,Cyclin-dependent kinase 2 ,Intracellular Signaling Peptides and Proteins ,Proteins ,Cell Biology ,Molecular biology ,COS Cells ,biology.protein ,Cyclin-dependent kinase 9 ,Protein Binding - Abstract
We have identified a novel serine/threonine kinase belonging to the myotonic dystrophy kinase family. The kinase can be produced in at least two different isoforms: a approximately 240-kDa protein (Citron Rho-interacting kinase, CRIK), in which the kinase domain is followed by the sequence of Citron, a previously identified Rho/Rac binding protein; a approximately 54-kDa protein (CRIK-short kinase (SK)), which consists mostly of the kinase domain. CRIK and CRIK-SK proteins are capable of phosphorylating exogenous substrates as well as of autophosphorylation, when tested by in vitro kinase assays after expression into COS7 cells. CRIK kinase activity is increased severalfold by coexpression of costitutively active Rho, while active Rac has more limited effects. Kinase activity of endogenous CRIK is indicated by in vitro kinase assays after immunoprecipitation with antibodies recognizing the Citron moiety of the protein. When expressed in keratinocytes, full-length CRIK, but not CRIK-SK, localizes into corpuscular cytoplasmic structures and elicits recruitment of actin into these structures. The previously reported Rho-associated kinases ROCK I and II are ubiquitously expressed. In contrast, CRIK exhibits a restricted pattern of expression, suggesting that this kinase may fulfill a more specialized function in specific cell types.
- Published
- 1998
13. STAT3-mediated activation of microRNA cluster 17 92 promotes proliferation and survival of ALK-positive anaplastic large cell lymphoma
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Spaccarotella, E., primary, Pellegrino, E., additional, Ferracin, M., additional, Ferreri, C., additional, Cuccuru, G., additional, Liu, C., additional, Iqbal, J., additional, Cantarella, D., additional, Taulli, R., additional, Provero, P., additional, Di Cunto, F., additional, Medico, E., additional, Negrini, M., additional, Chan, W. C., additional, Inghirami, G., additional, and Piva, R., additional
- Published
- 2013
- Full Text
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14. Evolution of Promoter Affinity for Transcription Factors in the Human Lineage
- Author
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Molineris, I., primary, Grassi, E., additional, Ala, U., additional, Di Cunto, F., additional, and Provero, P., additional
- Published
- 2011
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- View/download PDF
15. Structure of msj-1 gene in mice and humans: A possible role in the regulation of male reproduction
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Meccariello, R., primary, Berruti, G., additional, Chianese, R., additional, De Santis, R., additional, Di Cunto, F., additional, Scarpa, D., additional, Cobellis, G., additional, Zucchetti, I., additional, Pierantoni, R., additional, Altruda, F., additional, and Fasano, S., additional
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- 2008
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16. FINDING REGULATORY SITES FROM STATISTICAL ANALYSIS OF NUCLEOTIDE FREQUENCIES IN THE UPSTREAM REGION OF EUKARYOTIC GENES
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Caselle, M., primary, Provero, P., additional, Di Cunto, F., additional, and Pellegrino, M., additional
- Published
- 2002
- Full Text
- View/download PDF
17. Tissue-specific control of midbody microtubule stability by Citron kinase through modulation of TUBB3 phosphorylation
- Author
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Sgrò, F, Bianchi, F T, Falcone, M, Pallavicini, G, Gai, M, Chiotto, A M A, Berto, G E, Turco, E, Chang, Y J, Huttner, W B, and Di Cunto, F
- Abstract
Cytokinesis, the physical separation of daughter cells at the end of cell cycle, is commonly considered a highly stereotyped phenomenon. However, in some specialized cells this process may involve specific molecular events that are still largely unknown. In mammals, loss of Citron-kinase (CIT-K) leads to massive cytokinesis failure and apoptosis only in neuronal progenitors and in male germ cells, resulting in severe microcephaly and testicular hypoplasia, but the reasons for this specificity are unknown. In this report we show that CIT-K modulates the stability of midbody microtubules and that the expression of tubulin β-III (TUBB3) is crucial for this phenotype. We observed that TUBB3 is expressed in proliferating CNS progenitors, with a pattern correlating with the susceptibility to CIT-K loss. More importantly, depletion of TUBB3 in CIT-K-dependent cells makes them resistant to CIT-K loss, whereas TUBB3 overexpression increases their sensitivity to CIT-K knockdown. The loss of CIT-K leads to a strong decrease in the phosphorylation of S444 on TUBB3, a post-translational modification associated with microtubule stabilization. CIT-K may promote this event by interacting with TUBB3 and by recruiting at the midbody casein kinase-2α (CK2α) that has previously been reported to phosphorylate the S444 residue. Indeed, CK2α is lost from the midbody in CIT-K-depleted cells. Moreover, expression of the nonphosphorylatable TUBB3 mutant S444A induces cytokinesis failure, whereas expression of the phospho-mimetic mutant S444D rescues the cytokinesis failure induced by both CIT-K and CK2α loss. Altogether, our findings reveal that expression of relatively low levels of TUBB3 in mitotic cells can be detrimental for their cytokinesis and underscore the importance of CIT-K in counteracting this event.
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- 2016
- Full Text
- View/download PDF
18. The absence of p21Cip1/WAF1 alters keratinocyte growth and differentiation and promotes ras-tumor progression.
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Missero, C, primary, Di Cunto, F, additional, Kiyokawa, H, additional, Koff, A, additional, and Dotto, G P, additional
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- 1996
- Full Text
- View/download PDF
19. Ab initio identification of putative human transcription factor binding sites by comparative genomics.
- Author
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Corà, D., Herrmann, C., Dieterich, C., Di Cunto, F., Provero, P., and Caselle, M.
- Subjects
BINDING sites ,TRANSCRIPTION factors ,HUMAN genome ,GENE expression ,NUCLEOTIDE sequence ,BIOINFORMATICS - Abstract
Background: Understanding transcriptional regulation of gene expression is one of the greatest challenges of modern molecular biology. A central role in this mechanism is played by transcription factors, which typically bind to specific, short DNA sequence motifs usually located in the upstream region of the regulated genes. We discuss here a simple and powerful approach for the ab initio identification of these cis-regulatory motifs. The method we present integrates several elements: human-mouse comparison, statistical analysis of genomic sequences and the concept of coregulation. We apply it to a complete scan of the human genome. Results: By using the catalogue of conserved upstream sequences collected in the CORG database we construct sets of genes sharing the same overrepresented motif (short DNA sequence) in their upstream regions both in human and in mouse. We perform this construction for all possible motifs from 5 to 8 nucleotides in length and then filter the resulting sets looking for two types of evidence of coregulation: first, we analyze the Gene Ontology annotation of the genes in the set, searching for statistically significant common annotations; second, we analyze the expression profiles of the genes in the set as measured by microarray experiments, searching for evidence of coexpression. The sets which pass one or both filters are conjectured to contain a significant fraction of coregulated genes, and the upstream motifs characterizing the sets are thus good candidates to be the binding sites of the TF's involved in such regulation. In this way we find various known motifs and also some new candidate binding sites. Conclusion: We have discussed a new integrated algorithm for the "ab initio" identification of transcription factor binding sites in the human genome. The method is based on three ingredients: comparative genomics, over-representation, different types of coregulation. The method is applied to a full-scan of the human genome, giving satisfactory results. [ABSTRACT FROM AUTHOR]
- Published
- 2005
- Full Text
- View/download PDF
20. Citron rho-interacting kinase, a novel tissue-specific ser/thr kinase encompassing the Rho-Rac-binding protein Citron.
- Author
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Di Cunto, F, Calautti, E, Hsiao, J, Ong, L, Topley, G, Turco, E, and Dotto, G P
- Abstract
We have identified a novel serine/threonine kinase belonging to the myotonic dystrophy kinase family. The kinase can be produced in at least two different isoforms: a approximately 240-kDa protein (Citron Rho-interacting kinase, CRIK), in which the kinase domain is followed by the sequence of Citron, a previously identified Rho/Rac binding protein; a approximately 54-kDa protein (CRIK-short kinase (SK)), which consists mostly of the kinase domain. CRIK and CRIK-SK proteins are capable of phosphorylating exogenous substrates as well as of autophosphorylation, when tested by in vitro kinase assays after expression into COS7 cells. CRIK kinase activity is increased severalfold by coexpression of costitutively active Rho, while active Rac has more limited effects. Kinase activity of endogenous CRIK is indicated by in vitro kinase assays after immunoprecipitation with antibodies recognizing the Citron moiety of the protein. When expressed in keratinocytes, full-length CRIK, but not CRIK-SK, localizes into corpuscular cytoplasmic structures and elicits recruitment of actin into these structures. The previously reported Rho-associated kinases ROCK I and II are ubiquitously expressed. In contrast, CRIK exhibits a restricted pattern of expression, suggesting that this kinase may fulfill a more specialized function in specific cell types.
- Published
- 1998
21. p140Cap Regulates Memory and Synaptic Plasticity through Src-Mediated and Citron-N-Mediated Actin Reorganization
- Author
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Gaia Berto, Ferdinando Di Cunto, Nicoletta Berardi, Daniele Repetto, Tommaso Pizzorusso, Paola Di Stefano, Michela Matteoli, F. Bianchi, Markus Missler, Paola Camera, Paola Defilippi, Noemi Morello, Maurizio Giustetto, Isabella Russo, Eleonora Calcagno, Emilia Turco, Riccardo Melani, Romana Tomasoni, Repetto, D, Camera, P, Melani, R, Morello, N, Russo, I, Calcagno, E, Tomasoni, R, Bianchi, F, Berto, G, Giustetto, M, Berardi, N, Pizzorusso, T, Matteoli, M, Di Stefano, P, Missler, M, Turco, E, Di Cunto, F, and Defilippi, P.
- Subjects
Scaffold protein ,Patch-Clamp Techniques ,Dendritic spine ,spine dendritiche ,Dendritic Spines ,Blotting, Western ,Fluorescent Antibody Technique ,Protein Serine-Threonine Kinases ,recognition memory ,Hippocampus ,Synaptic Transmission ,Mice ,Actin remodeling of neurons ,Memory ,Animals ,Learning ,Mice, Knockout ,Neuronal Plasticity ,synaptic plasticity ,biology ,General Neuroscience ,Intracellular Signaling Peptides and Proteins ,Excitatory Postsynaptic Potentials ,perirhinal cortex ,Long-term potentiation ,Articles ,Actin cytoskeleton ,Actins ,Rats ,Cell biology ,Adaptor Proteins, Vesicular Transport ,src-Family Kinases ,Synaptic plasticity ,biology.protein ,Postsynaptic density ,Neuroscience ,Cortactin ,Signal Transduction - Abstract
A major challenge in the neuroscience field is the identification of molecules and pathways that control synaptic plasticity and memory. Dendritic spines play a pivotal role in these processes, as the major sites of excitatory synapses in neuronal communication. Previous studies have shown that the scaffold protein p140Cap localizes into dendritic spines and that its knockdown negatively modulates spine shape in culture. However, so far, there is no information on itsin vivorelevance. By using a knock-out mouse model, we here demonstrate that p140Cap is a key element for both learning and synaptic plasticity. Indeed,p140Cap−/−mice are impaired in object recognition test, as well as in LTP and in LTD measurements. Thein vivoeffects of p140Cap loss are presumably attenuated by noncell-autonomous events, since primary neurons obtained fromp140Cap−/−mice show a strong reduction in number of mushroom spines and abnormal organization of synapse-associated F-actin. These phenotypes are most likely caused by a local reduction of the inhibitory control of RhoA and of cortactin toward the actin-depolymerizing factor cofilin. These events can be controlled by p140Cap through its capability to directly inhibit the activation of Src kinase and by its binding to the scaffold protein Citron-N. Altogether, our results provide new insight into how protein associated with dynamic microtubules may regulate spine actin organization through interaction with postsynaptic density components.
- Published
- 2014
22. Identification of ΔNp63α Protein Interactions by Mass Spectrometry
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Viola Calabro, Angela Amoresano, Girolama La Mantia, Ferdinando Di Cunto, Antonella Di Costanzo, Gabriella Leo, Luisa Guerrini, Amoresano, Angela, DI COSTANZO, Antonella, Leo, Gabriella, Di Cunto, F., LA MANTIA, Girolama, Guerrini, L., and Calabro', Viola
- Subjects
Mass spectrometry ,integumentary system ,Chemistry ,protein-protein interactions ,P53 Tumor Suppressor ,Promoter ,General Chemistry ,Biochemistry ,Molecular biology ,Protein–protein interaction ,Cell biology ,Epithelial differentiation ,p53 gene family, mRNA metabolism ,Limb development ,Identification (biology) ,Gene ,Transcription factor - Abstract
p63, a transcription factor related to the p53 tumor suppressor, plays a key role in epidermal differentiation and limb development. The gene has two distinct promoters that allow the formation of proteins that either contain (TA) or lack (Delta N) a transactivation domain. Delta Np63 alpha is the most widely expressed isoform, at all stages of development and in adult tissues. It supports the regenerative capacity of basal keratinocytes and its upregulation is a hallmark of human squamous carcinomas. To get insight into the complex biology of Delta Np63 alpha, we set out to identify Delta Np63 alpha interacting proteins by co-immunoprecipitation in mammalian cells and mass spectrometry analysis. A total of 49 potential Delta Np63 alpha binding proteins, including several heterogeneous ribonucleoproteins (hnRNPs), were identified. Integration of the proteomic data with a Human Coexpression Network highlighted 5 putative p63 protein interactors whose expression is significantly comodulated with p63: hnRNPA/B, hnRNPK, hnRNPQ, FUS/TLS and Keratin 5. hnRNPA/B was already described as a p63 partner, but the others were novel. Interaction of Delta Np63 alpha with hnRNPQ, hnRNPK and FUS/TLS was confirmed by reciprocal co-immunoprecipitations in human keratinocytes. The finding that Delta Np63 alpha exists in complexes with several RNA-binding proteins lays the premises for the analysis of the role of Delta Np63 alpha in mRNA metabolism and transport.
- Published
- 2010
23. Structure of msj-1 gene in mice and humans: A possible role in the regulation of male reproduction
- Author
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Gilda Cobellis, Fiorella Altruda, F. Di Cunto, Donatella Scarpa, Ivana Zucchetti, Giovanna Berruti, Silvia Fasano, Riccardo Pierantoni, Rosaria Meccariello, Rosanna Chianese, R. De Santis, Meccariello, R, Berruti, G, Chianese, Rosanna, DE SANTIS, R, DI CUNTO, F, Scarpa, D, Cobellis, Gilda, Zucchetti, I, Altruda, F, Pierantoni, Riccardo, and Fasano, Silvia
- Subjects
Male ,DnaJ protein ,Mouse ,Molecular Sequence Data ,Inbred Strains ,Mice, Inbred Strains ,Nerve Tissue Proteins ,Biology ,DNAJ Protein ,Promoter Regions ,Mice ,Endocrinology ,Genetic ,Heat shock protein ,Testis ,Animals ,Humans ,Coding region ,Amino Acid Sequence ,msj-1 ,Promoter Regions, Genetic ,Gene ,Peptide sequence ,Genetics ,Base Sequence ,UDP glucuronosyltransferase ,Reproduction ,Intron ,Chromosome ,HSP40 Heat-Shock Proteins ,Spermatozoa ,Human ,Acrosome ,Molecular Chaperones ,Animal Science and Zoology ,Human genome - Abstract
Msj-1 gene encodes a DnaJ protein highly expressed in spermatids and spermatozoa of both rodents and amphibians, possibly involved in vesicle fusion and protein quality control by means of interaction with heat shock proteins. We isolated and characterized the entire murine msj -1 gene and searched for putative msj-1 -like genes into the human genome. Furthermore, ultrastructural localization of MSJ-1 was analyzed in mouse germ cells by immunogold electron microscopy. The analysis of murine msj-1 genomic sequence reveals that it is an intron less gene. Putative promoter region was predicted within the 600 bp upstream the transcription start site. In mouse, msj-1 maps on chromosome 1, into an intronic region of UDP glucuronosyl-transferase 1 family cluster. At ultrastructural level, MSJ-1 marks the developing acrosomic vesicle and the sperm centriolar region. A blast search against the human genome database revealed two closed regions ( Ha and Hb ) on human chromosome 2 having high nucleotide identity with murine msj-1 coding region. Similarly to mouse, in human both regions map into an intronic region of UDP glycosyl-transferase 1 family polypeptide A cluster ( ugt1a @). A significant ORF encoding a putative DnaJ protein of 145 aa was predicted from Ha . Finally, expression analysis, conducted by RT-PCR in human sperm cells, demonstrated that Ha mRNA is effectively present in humans; by Western blot, a specific MSJ-1 band of approximately 30 kDa was detected in human sperm. Taken together, these data suggest that msj-1 gene might be conserved among vertebrates and might exert fundamental functions in reproduction.
- Published
- 2008
24. Microarray and large-scale in silico-based identification of genes functionally related to Haptoglobin and/or Hemopexin
- Author
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Sharmila Fagoonee, Lorenzo Silengo, Emanuela Tolosano, Ferdinando Di Cunto, Paolo Gasparini, Diego Vozzi, Maurizio Pellegrino, Fiorella Altruda, Stefano Volinia, Fagoonee, S, DI CUNTO, F, Vozzi, Diego, Volinia, S, Pellegrino, M, Gasparini, Paolo, Silengo, L, Atruda, F, and Tolosano, E.
- Subjects
hemopexin ,Microarray ,In silico ,Computational biology ,Mice ,Complementary DNA ,Genetics ,polycyclic compounds ,Animals ,skin and connective tissue diseases ,Molecular Biology ,Gene ,Oligonucleotide Array Sequence Analysis ,Mice, Knockout ,Haptoglobins ,biology ,Reverse Transcriptase Polymerase Chain Reaction ,Microarray analysis techniques ,Haptoglobin ,Acute-phase protein ,food and beverages ,Hemopexin ,Cell Biology ,General Medicine ,haptoglobin ,Mice, Inbred C57BL ,body regions ,biology.protein ,microarray - Abstract
Haptoglobin and Hemopexin are plasma acute phase proteins that bind with high-affinity hemoglobin and heme, respectively. They play a key role in the protection against oxidative stress and inflammation. To dissect in more detail the mechanism of action of Haptoglobin and Hemopexin, it is important to identify their downstream effectors as well as genes functionally related to them. To this end, we performed a cDNA microarray analysis to compare gene expression profiles of the liver of Haptoglobin and Hemopexin single and double null mice to that of wild-type controls. Then, to extract the best candidates considered to be functionally related to Haptoglobin and/or Hemopexin from microarray-derived gene lists, we used a bioinformatic approach consisting in the screening of published microarray data for genes showing coexpression with Haptoglobin or Hemopexin. This strategy allowed us to identify a group of genes coexpressed with Haptoglobin or Hemopexin and transcriptionally modulated by their lack. These genes present a high probability to be functionally related to Haptoglobin and Hemopexin. Based on literature data, we picked up from this group of genes the ras suppressor Rsu1, the member of the G-protein signal transduction family Gnai2, and the cytokine Mdk as the best candidates mediating the anti-inflammatory action of Haptoglobin and Hemopexin.
- Published
- 2006
25. The absence of p21Cip1/WAF1 alters keratinocyte growth and differentiation and promotes ras-tumor progression
- Author
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F Di Cunto, Hiroaki Kiyokawa, Caterina Missero, Gian Paolo Dotto, Andrew Koff, Missero, Caterina, Di Cunto, F, Kiyokawa, H, Koff, A, and Dotto, Gp
- Subjects
Cyclin-Dependent Kinase Inhibitor p21 ,Keratinocytes ,Cellular differentiation ,Biology ,Mice ,Cyclin-dependent kinase ,Transforming Growth Factor beta ,Cyclins ,Genetics ,medicine ,Animals ,Mice, Knockout ,Oncogene ,Kinase ,Cell Differentiation ,Transforming growth factor beta ,Neoplasms, Experimental ,Cell biology ,medicine.anatomical_structure ,Cell Transformation, Neoplastic ,Genes, ras ,Tumor progression ,Knockout mouse ,biology.protein ,Calcium ,Keratinocyte ,Cell Division ,Developmental Biology - Abstract
p21Cip1/WAF1 was the first cyclin-dependent kinase (CDK) inhibitor to be identified, as a mediator of p53 in DNA damage-induced growth arrest, cell senescence, and direct CDK regulation. p21 may also play an important role in differentiation-associated growth arrest, as its expression is augmented in many terminally differentiating cells. A general involvement of p21 in growth/differentiation control and tumor suppression has been questioned, as mice lacking p21 undergo a normal development, harbor no gross alterations in any of their organs, and exhibit no increase in spontaneous tumor development. However, a significant imbalance between growth and differentiation could be unmasked under conditions where normal homeostatic mechanisms are impaired. We report here that primary keratinocytes derived from p21 knockout mice, transformed with a ras oncogene, and injected subcutaneously into nude mice exhibit a very aggressive tumorigenic behavior, which is not observed with wild-type control keratinocytes nor with keratinocytes with a disruption of the closely related p27 gene. p21 knockout keratinocytes tested under well-defined in vitro conditions show a significantly increased proliferative potential, which is also observed but to a lesser extent with p27 knockout cells. More profound differences were found in the differentiation behavior of p21 versus p27 knockout keratinocytes, with p21 (but not p27) deficiency causing a drastic down-modulation of differentiation markers linked with the late stages of the keratinocyte terminal differentiation program. Thus, our results reveal a so far undetected role of p21 in tumor suppression, demonstrate that this function is specific as it cannot be attributed to the closely related p27 molecule, and point to an essential involvement of p21 in terminal differentiation control, which may account for its role in tumor suppression.
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- 1996
26. Modeling primary microcephaly with human brain organoids reveals fundamental roles of CIT kinase activity.
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Pallavicini G, Moccia A, Iegiani G, Parolisi R, Peirent ER, Berto GE, Lorenzati M, Tshuva RY, Ferraro A, Balzac F, Turco E, Salvi SU, Myklebust HF, Wang S, Eisenberg J, Chitale M, Girgla NS, Boda E, Reiner O, Buffo A, Di Cunto F, and Bielas SL
- Abstract
Brain size and cellular heterogeneity are tightly regulated by species-specific proliferation and differentiation of multipotent neural progenitor cells (NPCs). Errors in this process are among the mechanisms of primary hereditary microcephaly (MCPH), a group of disorders characterized by reduced brain size and intellectual disability. Biallelic CIT missense variants that disrupt kinase function (CITKI/KI) and frameshift loss-of-function variants (CITFS/FS) are the genetic basis for MCPH17; however, the function of CIT catalytic activity in brain development and NPC cytokinesis is unknown. Therefore, we created the CitKI/KI mouse model and found that it does not phenocopy human microcephaly, unlike biallelic CitFS/FS animals. Nevertheless, both Cit models exhibited binucleation, DNA damage, and apoptosis. To investigate human-specific mechanisms of CIT microcephaly, we generated CITKI/KI and CITFS/FS human forebrain organoids. We found that CITKI/KI and CITFS/FS organoids lose cytoarchitectural complexity, transitioning from pseudostratified to simple neuroepithelium. This change was associated with defects that disrupt polarity of NPC cytokinesis, in addition to elevating apoptosis. Together, our results indicate that both CIT catalytic and scaffolding functions in NPC cytokinesis are critical for human corticogenesis. Species differences in corticogenesis and the dynamic 3D features of NPC mitosis underscore the utility of human forebrain organoid models for understanding human microcephaly.
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- 2024
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27. Standardization of a Novel Semi-Automatic Software for Neurite Outgrowth Measurement.
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Musso G, Dotta S, Parmar A, Rasà DM, Di Cunto F, and Marvaldi L
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- Animals, Neurites, Algorithms, Mice, Image Processing, Computer-Assisted methods, Rats, Software, Neuronal Outgrowth physiology
- Abstract
Effective live-imaging techniques are crucial to assess neuronal morphology in order to measure neurite outgrowth in real time. The proper measurement of neurite outgrowth has been a long-standing challenge over the years in the neuroscience research field. This parameter serves as a cornerstone in numerous in vitro experimental setups, ranging from dissociated cultures and organotypic cultures to cell lines. By quantifying the neurite length, it is possible to determine if a specific treatment worked or if axonal regeneration is enhanced in different experimental groups. In this study, the aim is to demonstrate the robustness and accuracy of the Incucyte Neurotrack neurite outgrowth analysis software. This semi-automatic software is available in a time-lapse microscopy system which offers several advantages over commonly used methodologies in the quantification of the neurite length in phase contrast images. The algorithm masks and quantifies several parameters in each image and returns neuronal cell metrics, including neurite length, branch points, cell-body clusters, and cell-body cluster areas. Firstly, we validated the robustness and accuracy of the software by correlating its values with those of the manual NeuronJ, a Fiji plug-in. Secondly, we used the algorithm which is able to work both on phase contrast images as well as on immunocytochemistry images. Using specific neuronal markers, we validated the feasibility of the fluorescence-based neurite outgrowth analysis on sensory neurons in vitro cultures. Additionally, this software can measure neurite length across various seeding conditions, ranging from individual cells to complex neuronal nets. In conclusion, the software provides an innovative and time-effective platform for neurite outgrowth assays, paving the way for faster and more reliable quantifications.
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- 2024
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28. Correction: The Down syndrome critical region protein TTC3 inhibits neuronal differentiation via RhoA and Citron kinase.
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Berto G, Camera P, Fusco C, Imarisio S, Ambrogio C, Chiarle R, Silengo L, and Di Cunto F
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- 2023
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29. The impact of TP53 activation and apoptosis in primary hereditary microcephaly.
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Iegiani G, Ferraro A, Pallavicini G, and Di Cunto F
- Abstract
Autosomal recessive primary microcephaly (MCPH) is a constellation of disorders that share significant brain size reduction and mild to moderate intellectual disability, which may be accompanied by a large variety of more invalidating clinical signs. Extensive neural progenitor cells (NPC) proliferation and differentiation are essential to determine brain final size. Accordingly, the 30 MCPH loci mapped so far (MCPH1-MCPH30) encode for proteins involved in microtubule and spindle organization, centriole biogenesis, nuclear envelope, DNA replication and repair, underscoring that a wide variety of cellular processes is required for sustaining NPC expansion during development. Current models propose that altered balance between symmetric and asymmetric division, as well as premature differentiation, are the main mechanisms leading to MCPH. Although studies of cellular alterations in microcephaly models have constantly shown the co-existence of high DNA damage and apoptosis levels, these mechanisms are less considered as primary factors. In this review we highlight how the molecular and cellular events produced by mutation of the majority of MCPH genes may converge on apoptotic death of NPCs and neurons, via TP53 activation. We propose that these mechanisms should be more carefully considered in the alterations of the sophisticated equilibrium between proliferation, differentiation and death produced by MCPH gene mutations. In consideration of the potential druggability of cell apoptotic pathways, a better understanding of their role in MCPH may significantly facilitate the development of translational approaches., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Iegiani, Ferraro, Pallavicini and Di Cunto.)
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- 2023
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30. Lestaurtinib inhibits Citron kinase activity and medulloblastoma growth through induction of DNA damage, apoptosis and cytokinesis failure.
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Pallavicini G, Iegiani G, Parolisi R, Ferraro A, Garello F, Bitonto V, Terreno E, Gai M, and Di Cunto F
- Abstract
Introduction: Medulloblastoma (MB), the most common malignant pediatric brain tumor, is currently treated with surgery followed by radiation and chemotherapy, which is accompanied by severe side effects, raising the need for innovative therapies. Disruption of the microcephaly-related gene Citron kinase (CITK) impairs the expansion of xenograft models as well as spontaneous MB arising in transgenic mice. No specific CITK inhibitors are available., Methods: Lestaurtinib, a Staurosporine derivative also known as CEP-701, inhibits CITK with IC50 of 90 nM. We therefore tested the biological effects of this molecule on different MB cell lines, as well as in vivo, injecting the drug in MBs arising in SmoA1 transgenic mice., Results: Similar to CITK knockdown, treatment of MB cells with 100 nM Lestaurtinib reduces phospho-INCENP levels at the midbody and leads to late cytokinesis failure. Moreover, Lestaurtinib impairs cell proliferation through CITK-sensitive mechanisms. These phenotypes are accompanied by accumulation of DNA double strand breaks, cell cycle block and TP53 superfamily activation in vitro and in vivo. Lestaurtinib treatment reduces tumor growth and increases mice survival., Discussion: Our data indicate that Lestaurtinib produces in MB cells poly-pharmacological effects extending beyond the inhibition of its validated targets, supporting the possibility of repositioning this drug for MB treatment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Pallavicini, Iegiani, Parolisi, Ferraro, Garello, Bitonto, Terreno, Gai and Di Cunto.)
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- 2023
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31. Molecular and functional heterogeneity in dorsal and ventral oligodendrocyte progenitor cells of the mouse forebrain in response to DNA damage.
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Boda E, Lorenzati M, Parolisi R, Harding B, Pallavicini G, Bonfanti L, Moccia A, Bielas S, Di Cunto F, and Buffo A
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- Animals, DNA Damage, Mice, Oligodendroglia metabolism, Prosencephalon, Oligodendrocyte Precursor Cells physiology
- Abstract
In the developing mouse forebrain, temporally distinct waves of oligodendrocyte progenitor cells (OPCs) arise from different germinal zones and eventually populate either dorsal or ventral regions, where they present as transcriptionally and functionally equivalent cells. Despite that, developmental heterogeneity influences adult OPC responses upon demyelination. Here we show that accumulation of DNA damage due to ablation of citron-kinase or cisplatin treatment cell-autonomously disrupts OPC fate, resulting in cell death and senescence in the dorsal and ventral subsets, respectively. Such alternative fates are associated with distinct developmental origins of OPCs, and with a different activation of NRF2-mediated anti-oxidant responses. These data indicate that, upon injury, dorsal and ventral OPC subsets show functional and molecular diversity that can make them differentially vulnerable to pathological conditions associated with DNA damage., (© 2022. The Author(s).)
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- 2022
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32. Inhibiting microcephaly genes as alternative to microtubule targeting agents to treat brain tumors.
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Iegiani G, Di Cunto F, and Pallavicini G
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- Animals, Apoptosis, Brain Neoplasms pathology, Clinical Trials as Topic, Cytokinesis, Humans, Microcephaly pathology, Brain Neoplasms genetics, Brain Neoplasms therapy, Microcephaly genetics, Microtubules metabolism
- Abstract
Medulloblastoma (MB) and gliomas are the most frequent high-grade brain tumors (HGBT) in children and adulthood, respectively. The general treatment for these tumors consists in surgery, followed by radiotherapy and chemotherapy. Despite the improvement in patient survival, these therapies are only partially effective, and many patients still die. In the last decades, microtubules have emerged as interesting molecular targets for HGBT, as various microtubule targeting agents (MTAs) have been developed and tested pre-clinically and clinically with encouraging results. Nevertheless, these treatments produce relevant side effects since they target microtubules in normal as well as in cancerous cells. A possible strategy to overcome this toxicity could be to target proteins that control microtubule dynamics but are required by HGBT cells much more than in normal cell types. The genes mutated in primary hereditary microcephaly (MCPH) are ubiquitously expressed in proliferating cells, but under normal conditions are selectively required during brain development, in neural progenitors. There is evidence that MB and glioma cells share molecular profiles with progenitors of cerebellar granules and of cortical radial glia cells, in which MCPH gene functions are fundamental. Moreover, several studies indicate that MCPH genes are required for HGBT expansion. Among the 25 known MCPH genes, we focus this review on KNL1, ASPM, CENPE, CITK and KIF14, which have been found to control microtubule stability during cell division. We summarize the current knowledge about the molecular basis of their interaction with microtubules. Moreover, we will discuss data that suggest these genes are promising candidates as HGBT-specific targets., (© 2021. The Author(s).)
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- 2021
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33. Goldberg-Shprintzen syndrome protein KIF1BP is a CITK interactor implicated in cytokinesis.
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Pallavicini G, Gai M, Iegiani G, Berto GE, Adrait A, Couté Y, and Di Cunto F
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- Cytokinesis genetics, HeLa Cells, Humans, Spindle Apparatus, Craniofacial Abnormalities, Hirschsprung Disease
- Abstract
Goldberg-Shprintzen disease (GOSHS) is a rare microcephaly syndrome accompanied by intellectual disability, dysmorphic facial features, peripheral neuropathy and Hirschsprung disease. It is associated with recessive mutations in the gene encoding kinesin family member 1-binding protein (KIF1BP, also known as KIFBP). The encoded protein regulates axon microtubules dynamics, kinesin attachment and mitochondrial biogenesis, but it is not clear how its loss could lead to microcephaly. We identified KIF1BP in the interactome of citron kinase (CITK, also known as CIT), a protein produced by the primary hereditary microcephaly 17 (MCPH17) gene. KIF1BP and CITK interact under physiological conditions in mitotic cells. Similar to CITK, KIF1BP is enriched at the midbody ring and is required for cytokinesis. The association between KIF1BP and CITK can be influenced by CITK activity, and the two proteins may antagonize each other for their midbody localization. KIF1BP knockdown decreases microtubule stability, increases KIF23 midbody levels and impairs midbody localization of KIF14, as well as of chromosome passenger complex. These data indicate that KIF1BP is a CITK interactor involved in midbody maturation and abscission, and suggest that cytokinesis failure may contribute to the microcephaly phenotype observed in GOSHS., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)
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- 2021
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34. miR-7b-3p Exerts a Dual Role After Spinal Cord Injury, by Supporting Plasticity and Neuroprotection at Cortical Level.
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Ghibaudi M, Boido M, Green D, Signorino E, Berto GE, Pourshayesteh S, Singh A, Di Cunto F, Dalmay T, and Vercelli A
- Abstract
Spinal cord injury (SCI) affects 6 million people worldwide with no available treatment. Despite research advances, the inherent poor regeneration potential of the central nervous system remains a major hurdle. Small RNAs (sRNAs) 19-33 nucleotides in length are a set of non-coding RNA molecules that regulate gene expression and have emerged as key players in regulating cellular events occurring after SCI. Here we profiled a class of sRNA known as microRNAs (miRNAs) following SCI in the cortex where the cell bodies of corticospinal motor neurons are located. We identified miR-7b-3p as a candidate target given its significant upregulation after SCI in vivo and we screened by miRWalk PTM the genes predicted to be targets of miR-7b-3p (among which we identified Wipf2 , a gene regulating neurite extension). Moreover, 16 genes, involved in neural regeneration and potential miR-7b-3p targets, were found to be downregulated in the cortex following SCI. We also analysed miR-7b-3p function during cortical neuron development in vitro : we observed that the overexpression of miR-7b-3p was important (1) to maintain neurons in a more immature and, likely, plastic neuronal developmental phase and (2) to contrast the apoptotic pathway; however, in normal conditions it did not affect the Wipf2 expression. On the contrary, the overexpression of miR-7b-3p upon in vitro oxidative stress condition (mimicking the SCI environment) significantly reduced the expression level of Wipf2, as observed in vivo , confirming it as a direct miR-7b-3p target. Overall, these data suggest a dual role of miR-7b-3p: (i) the induction of a more plastic neuronal condition/phase, possibly at the expense of the axon growth, (ii) the neuroprotective role exerted through the inhibition of the apoptotic cascade. Increasing the miR-7b-3p levels in case of SCI could reactivate in adult neurons silenced developmental programmes, supporting at the same time the survival of the axotomised neurons., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ghibaudi, Boido, Green, Signorino, Berto, Pourshayesteh, Singh, Di Cunto, Dalmay and Vercelli.)
- Published
- 2021
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35. CENPE Inhibition Leads to Mitotic Catastrophe and DNA Damage in Medulloblastoma Cells.
- Author
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Iegiani G, Gai M, Di Cunto F, and Pallavicini G
- Abstract
Medulloblastoma (MB) is the most frequent brain tumor in children. The standard treatment consists in surgery, followed by radiotherapy and chemotherapy. These therapies are only partially effective since many patients still die and those who survive suffer from neurological and endocrine disorders. Therefore, more effective therapies are needed. Primary microcephaly (MCPH) is a rare disorder caused by mutations in 25 different genes. Centromere-associated protein E (CENPE) heterozygous mutations cause the MCPH13 syndrome. As for other MCPH genes, CENPE is required for normal proliferation and survival of neural progenitors. Since there is evidence that MB shares many molecular features with neural progenitors, we hypothesized that CENPE could be an effective target for MB treatment. In ONS-76 and DAOY cells, CENPE knockdown induced mitotic defects and apoptosis. Moreover, CENPE depletion induced endogenous DNA damage accumulation, activating TP53 or TP73 as well as cell death signaling pathways. To consolidate CENPE as a target for MB treatment, we tested GSK923295, an allosteric inhibitor already in clinical trial for other cancer types. GSK923295, induced effects similar to CENPE depletion with higher penetrance, at low nM levels, suggesting that CENPE's inhibition could be a therapeutic strategy for MB treatment.
- Published
- 2021
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36. MECP2 mutations affect ciliogenesis: a novel perspective for Rett syndrome and related disorders.
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Frasca A, Spiombi E, Palmieri M, Albizzati E, Valente MM, Bergo A, Leva B, Kilstrup-Nielsen C, Bianchi F, Di Carlo V, Di Cunto F, and Landsberger N
- Subjects
- Animals, Brain metabolism, Hedgehog Proteins, Humans, Methyl-CpG-Binding Protein 2 genetics, Methyl-CpG-Binding Protein 2 metabolism, Mice, Mutation, Rett Syndrome genetics
- Abstract
Mutations in MECP2 cause several neurological disorders of which Rett syndrome (RTT) represents the best-defined condition. Although mainly working as a transcriptional repressor, MeCP2 is a multifunctional protein revealing several activities, the involvement of which in RTT remains obscure. Besides being mainly localized in the nucleus, MeCP2 associates with the centrosome, an organelle from which primary cilia originate. Primary cilia function as "sensory antennae" protruding from most cells, and a link between primary cilia and mental illness has recently been reported. We herein demonstrate that MeCP2 deficiency affects ciliogenesis in cultured cells, including neurons and RTT fibroblasts, and in the mouse brain. Consequently, the cilium-related Sonic Hedgehog pathway, which is essential for brain development and functioning, is impaired. Microtubule instability participates in these phenotypes that can be rescued by HDAC6 inhibition together with the recovery of RTT-related neuronal defects. Our data indicate defects of primary cilium as a novel pathogenic mechanism that by contributing to the clinical features of RTT might impact on proper cerebellum/brain development and functioning, thus providing a novel therapeutic target., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)
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- 2020
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37. Correction to: The SRCIN1/p140Cap adaptor protein negatively regulates the aggressiveness of neuroblastoma.
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Grasso S, Cangelosi D, Chapelle J, Alzona M, Centonze G, Lamolinara A, Salemme V, Angelini C, Morellato A, Saglietto A, Bianchi FT, Cabodi S, Salaroglio IC, Fusella F, Ognibene M, Iezzi M, Pezzolo A, Poli V, Di Cunto F, Eva A, Riganti C, Varesio L, Turco E, and Defilippi P
- Abstract
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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- 2020
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38. Of rings and spines: The multiple facets of Citron proteins in neural development.
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Bianchi FT, Gai M, Berto GE, and Di Cunto F
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- Animals, Gene Expression Regulation, Enzymologic, Genetic Variation, Humans, Intracellular Signaling Peptides and Proteins genetics, Nervous System cytology, Neurons cytology, Phenotype, Protein Serine-Threonine Kinases genetics, Intracellular Signaling Peptides and Proteins metabolism, Nervous System growth & development, Protein Serine-Threonine Kinases metabolism
- Abstract
The Citron protein was originally identified for its capability to specifically bind the active form of RhoA small GTPase, leading to the simplistic hypothesis that it may work as a RhoA downstream effector in actin remodeling. More than two decades later, a much more complex picture has emerged. In particular, it has become clear that in animals, and especially in mammals, the functions of the Citron gene ( CIT ) are intimately linked to many aspects of central nervous system (CNS) development and function, although the gene is broadly expressed. More specifically, CIT encodes two main isoforms, Citron-kinase (CIT-K) and Citron-N (CIT-N), characterized by complementary expression pattern and different functions. Moreover, in many of their activities, CIT proteins act more as upstream regulators than as downstream effectors of RhoA. Finally it has been found that, besides working through actin, CIT proteins have many crucial functional interactions with the microtubule cytoskeleton and may directly affect genome stability. In this review, we will summarize these advances and illustrate their actual or potential relevance for CNS diseases, including microcephaly and psychiatric disorders.
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- 2020
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39. CITK Loss Inhibits Growth of Group 3 and Group 4 Medulloblastoma Cells and Sensitizes Them to DNA-Damaging Agents.
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Pallavicini G, Iegiani G, Berto GE, Calamia E, Trevisiol E, Veltri A, Allis S, and Di Cunto F
- Abstract
Medulloblastoma (MB) is the most common malignant brain tumor in children, and it is classified into four biological subgroups: WNT, Sonic Hedgehog (SHH), Group 3 and Group 4. The current treatment is surgery, followed by irradiation and chemotherapy. Unfortunately, these therapies are only partially effective. Citron kinase protein (CITK) has been proposed as a promising target for SHH MB, whose inactivation leads to DNA damage and apoptosis. D283 and D341 cell lines (Group 3/Group 4 MB) were silenced with established siRNA sequences against CITK, to assess the direct effects of its loss. Next, D283, D341, ONS-76 and DAOY cells were treated with ionizing radiation (IR) or cisplatin in combination with CITK knockdown. CITK depletion impaired proliferation and induced cytokinesis failure and apoptosis of G3/G4 MB cell lines. Furthermore, CITK knockdown produced an accumulation of DNA damage, with reduced RAD51 nuclear levels. Association of IR or cisplatin with CITK depletion strongly impaired the growth potential of all tested MB cells. These results indicate that CITK inactivation could prevent the expansion of G3/G4 MB and increase their sensitivity to DNA-damaging agents, by impairing homologous recombination. We suggest that CITK inhibition could be broadly associated with IR and adjuvant therapy in MB treatment., Competing Interests: The authors declare no conflicts of interest.
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- 2020
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40. The SRCIN1/p140Cap adaptor protein negatively regulates the aggressiveness of neuroblastoma.
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Grasso S, Cangelosi D, Chapelle J, Alzona M, Centonze G, Lamolinara A, Salemme V, Angelini C, Morellato A, Saglietto A, Bianchi FT, Cabodi S, Salaroglio IC, Fusella F, Ognibene M, Iezzi M, Pezzolo A, Poli V, Di Cunto F, Eva A, Riganti C, Varesio L, Turco E, and Defilippi P
- Subjects
- Adaptor Proteins, Vesicular Transport genetics, Animals, Biomarkers, Tumor genetics, Cell Proliferation, Cell Survival, Humans, Infant, Lung Neoplasms diagnosis, Lung Neoplasms metabolism, Male, Mice, Mice, Inbred NOD, Mice, SCID, Neoplasms, Experimental metabolism, Neoplasms, Experimental pathology, Neuroblastoma diagnosis, RNA, Messenger genetics, RNA, Messenger metabolism, Tumor Cells, Cultured, Adaptor Proteins, Vesicular Transport metabolism, Biomarkers, Tumor metabolism, Lung Neoplasms secondary, Neuroblastoma metabolism
- Abstract
Neuroblastoma is the most common extra-cranial pediatric solid tumor, responsible for 13-15% of pediatric cancer death. Its intrinsic heterogeneity makes it difficult to target for successful therapy. The adaptor protein p140Cap/SRCIN1 negatively regulates tumor cell features and limits breast cancer progression. This study wish to assess if p140Cap is a key biological determinant of neuroblastoma outcome. RNAseq profiles of a large cohort of neuroblastoma patients show that SRCIN1 mRNA levels are an independent risk factor inversely correlated to disease aggressiveness. In high-risk patients, CGH+SNP microarray analysis of primary neuroblastoma identifies SRCIN1 as frequently altered by hemizygous deletion, copy-neutral loss of heterozygosity, or disruption. Functional experiments show that p140Cap negatively regulates Src and STAT3 signaling, affects anchorage-independent growth and migration, in vivo tumor growth and spontaneous lung metastasis formation. p140Cap also increases sensitivity of neuroblastoma cells to doxorubicin and etoposide treatment, as well as to a combined treatment with chemotherapy drugs and Src inhibitors. Our functional findings point to a causal role of p140Cap in curbing the aggressiveness of neuroblastoma, due to its ability to impinge on specific molecular pathways, and to sensitize cells to therapeutic treatment. This study provides the first evidence that the SRCIN1/p140Cap adaptor protein is a key player in neuroblastoma as a new independent prognostic marker for patient outcome and treatment. Altogether, these data highlight the potential clinical impact of SRCIN1/p140Cap expression in neuroblastoma tumors, in terms of reducing cytotoxic effects of chemotherapy, one of the main issues for pediatric tumor treatment.
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- 2020
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41. Citron kinase interacts with LATS2 and inhibits its activity by occluding its hydrophobic phosphorylation motif.
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Tran THY, Yang DW, Kim M, Lee DH, Gai M, Di Cunto F, Choi KW, and Lim DS
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- Amino Acid Motifs, Animals, Drosophila Proteins metabolism, Drosophila melanogaster metabolism, Epistasis, Genetic, Genotype, Humans, Hydrophobic and Hydrophilic Interactions, Intracellular Signaling Peptides and Proteins genetics, Models, Biological, Nuclear Proteins metabolism, Phenotype, Phosphorylation, Protein Binding, Protein Serine-Threonine Kinases chemistry, Protein Serine-Threonine Kinases genetics, Trans-Activators metabolism, YAP-Signaling Proteins, Intracellular Signaling Peptides and Proteins chemistry, Intracellular Signaling Peptides and Proteins metabolism, Protein Interaction Domains and Motifs, Protein Serine-Threonine Kinases antagonists & inhibitors, Protein Serine-Threonine Kinases metabolism
- Abstract
The inhibitory effect of large tumor suppressor kinase (LATS1/2) on the activity of the oncoprotein yes-associated protein (YAP) is crucial to maintain tissue homeostasis. Proteomic studies have identified several new regulators of this process. Recently, citron kinase (CIT) was listed as a potential binding candidate of Hippo-related components, suggesting a new connection between CIT and the Hippo pathway. Aside from CIT's role in cytokinesis, the molecular crosstalk between CIT and the Hippo pathway is largely unknown. Here, we demonstrate a role for CIT as a scaffold protein linking LATS2 and YAP. More importantly, CIT interacts with LATS2 to directly suppress LATS2 phosphorylation at the hydrophobic motif-targeted by MST1, leading to LATS2 inactivation and YAP activation. By studying their genetic interactions, we found that Sticky, the CIT homolog in Drosophila melanogaster, functions with Warts to control Drosophila eye development. Together, our study confirms citron kinase as a novel regulator of the Hippo pathway., (© The Author(s) (2019). Published by Oxford University Press on behalf of Journal of Molecular Cell Biology, IBCB, SIBS, CAS.)
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- 2019
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42. Neuronal Cell-Intrinsic Defects in Mouse Models of Down Syndrome.
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Chiotto AMA, Migliorero M, Pallavicini G, Bianchi FT, Gai M, Di Cunto F, and Berto GE
- Abstract
Down Syndrome (DS) is the most common genetic disorder associated with intellectual disability (ID). Excitatory neurons of DS patients and mouse models show decreased size of dendritic field and reduction of spine density. Whether these defects are caused by cell autonomous alterations or by abnormal multicellular circuitry is still unknown. In this work, we explored this issue by culturing cortical neurons obtained from two mouse models of DS: the widely used Ts65Dn and the less characterized Ts2Cje. We observed that, in the in vitro conditions, axon specification and elongation, as well as dendritogenesis, take place without evident abnormalities, indicating that the initial phases of neuronal differentiation do not suffer from the presence of an imbalanced genetic dosage. Conversely, our analysis highlighted differences between trisomic and euploid neurons in terms of reduction of spine density, in accordance with in vivo data obtained by other groups, proposing the presence of a cell-intrinsic malfunction. This work suggests that the characteristic morphological defects of DS neurons are likely to be caused by the possible combination of cell-intrinsic defects together with cell-extrinsic cues. Additionally, our data support the possibility of using the more sustainable line Ts2Cje as a standard model for the study of DS., (Copyright © 2019 Chiotto, Migliorero, Pallavicini, Bianchi, Gai, Di Cunto and Berto.)
- Published
- 2019
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43. Precision Revisited: Targeting Microcephaly Kinases in Brain Tumors.
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Pallavicini G, Berto GE, and Di Cunto F
- Subjects
- Animals, Glioblastoma metabolism, Glioblastoma pathology, Humans, Medulloblastoma metabolism, Medulloblastoma pathology, Protein Serine-Threonine Kinases metabolism, Brain Neoplasms metabolism, Brain Neoplasms pathology, Microcephaly metabolism, Microcephaly pathology
- Abstract
Glioblastoma multiforme and medulloblastoma are the most frequent high-grade brain tumors in adults and children, respectively. Standard therapies for these cancers are mainly based on surgical resection, radiotherapy, and chemotherapy. However, intrinsic or acquired resistance to treatment occurs almost invariably in the first case, and side effects are unacceptable in the second. Therefore, the development of new, effective drugs is a very important unmet medical need. A critical requirement for developing such agents is to identify druggable targets required for the proliferation or survival of tumor cells, but not of other cell types. Under this perspective, genes mutated in congenital microcephaly represent interesting candidates. Congenital microcephaly comprises a heterogeneous group of disorders in which brain volume is reduced, in the absence or presence of variable syndromic features. Genetic studies have clarified that most microcephaly genes encode ubiquitous proteins involved in mitosis and in maintenance of genomic stability, but the effects of their inactivation are particularly strong in neural progenitors. It is therefore conceivable that the inhibition of the function of these genes may specifically affect the proliferation and survival of brain tumor cells. Microcephaly genes encode for a few kinases, including CITK, PLK4, AKT3, DYRK1A, and TRIO. In this review, we summarize the evidence indicating that the inhibition of these molecules could exert beneficial effects on different aspects of brain cancer treatment., Competing Interests: The authors declare no conflict of interest.
- Published
- 2019
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44. A novel p.Ser108LeufsTer15 SOD1 mutation leading to the formation of a premature stop codon in an apparently sporadic ALS patient: insights into the underlying pathomechanisms.
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Canosa A, De Marco G, Lomartire A, Rinaudo MT, Di Cunto F, Turco E, Barberis M, Brunetti M, Casale F, Moglia C, Calvo A, Marklund SL, Andersen PM, Mora G, and Chiò A
- Subjects
- Aged, Female, Frameshift Mutation, Humans, Amyotrophic Lateral Sclerosis diagnosis, Amyotrophic Lateral Sclerosis genetics, Superoxide Dismutase-1 genetics
- Abstract
We report an apparently sporadic amyotrophic lateral sclerosis patient carrying a heterozygous novel frameshift SOD1 mutation (p.Ser108LeufsTer15), predicted to cause a premature protein truncation. RT-PCR analysis of SOD1 mRNA and SDS-PAGE/Western blot analysis of PBMC demonstrated that mRNA from the mutant allele is expressed at levels similar to those of the wild-type allele, but the truncated protein is undetectable also in the insoluble fraction and after proteasome inhibition. Accordingly, the dismutation activity in erythrocytes is halved. Thus, the pathogenic mechanism associated with this mutation might be based on an insufficient activity of SOD1 that would make motor neurons more vulnerable to oxidative injury. However, it cannot be excluded that p.Ser108LeufsTer15 SOD1 is present in the nervous tissue and, being less charged and hence having less repulsive forces than the wild-type protein, may trigger toxic mechanisms as a consequence of its propensity to aggregate., (Copyright © 2018 Elsevier Inc. All rights reserved.)
- Published
- 2018
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45. p53-Sensitive Epileptic Behavior and Inflammation in Ft1 Hypomorphic Mice.
- Author
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Burla R, La Torre M, Zanetti G, Bastianelli A, Merigliano C, Del Giudice S, Vercelli A, Di Cunto F, Boido M, Vernì F, and Saggio I
- Abstract
Epilepsy is a complex clinical condition characterized by repeated spontaneous seizures. Seizures have been linked to multiple drivers including DNA damage accumulation. Investigation of epilepsy physiopathology in humans imposes ethical and practical limitations, for this reason model systems are mostly preferred. Among animal models, mouse mutants are particularly valuable since they allow conjoint behavioral, organismal, and genetic analyses. Along with this, since aging has been associated with higher frequency of seizures, prematurely aging mice, simulating human progeroid diseases, offer a further useful modeling element as they recapitulate aging over a short time-window. Here we report on a mouse mutant with progeroid traits that displays repeated spontaneous seizures. Mutant mice were produced by reducing the expression of the gene Ft1 ( AKTIP in humans). In vitro , AKTIP/Ft1 depletion causes telomere aberrations, DNA damage, and cell senescence. AKTIP/Ft1 interacts with lamins, which control nuclear architecture and DNA function. Premature aging defects of Ft1 mutant mice include skeletal alterations and lipodystrophy. The epileptic behavior of Ft1 mutant animals was age and sex linked. Seizures were observed in 18 mutant mice (23.6% of aged ≥ 21 weeks), at an average frequency of 2.33 events/mouse. Time distribution of seizures indicated non-random enrichment of seizures over the follow-up period, with 75% of seizures happening in consecutive weeks. The analysis of epileptic brains did not reveal overt brain morphological alterations or severe neurodegeneration, however, Ft1 reduction induced expression of the inflammatory markers IL-6 and TGF-β. Importantly, Ft1 mutant mice with concomitant genetic reduction of the guardian of the genome, p53, showed no seizures or inflammatory marker activation, implicating the DNA damage response into these phenotypes. This work adds insights into the connection among DNA damage, brain function, and aging. In addition, it further underscores the importance of model organisms for studying specific phenotypes, along with permitting the analysis of genetic interactions at the organismal level.
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- 2018
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46. Correction to: ZIKA virus elicits P53 activation and genotoxic stress in human neural progenitors similar to mutations involved in severe forms of genetic microcephaly.
- Author
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El Ghouzzi V, Bianchi FT, Molineris I, Mounce BC, Berto GE, Rak M, Lebon S, Aubry L, Tocco C, Gai M, Chiotto AMA, Sgrò F, Pallavicini G, Simon-Loriere E, Passemard S, Vignuzzi M, Gressens P, and Di Cunto F
- Abstract
The authors wish to point out that the name of the first author is appearing incorrectly on Pubmed: it should be El Ghouzzi V (and not Ghouzzi VE). In addition, the words "and p53" appear at the end of the title in the original publication ( https://www.nature.com/articles/cddis2016266 ) and in the previous erratum version ( https://www.nature.com/articles/cddis2016446 ). This is not correct.
- Published
- 2018
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47. Impact of DNA repair and stability defects on cortical development.
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Bianchi FT, Berto GE, and Di Cunto F
- Subjects
- Fanconi Anemia pathology, Humans, Neurogenesis genetics, DNA Damage genetics, DNA Repair genetics, Fanconi Anemia genetics, Genomic Instability genetics
- Abstract
Maintenance of genome stability is a crucial cellular function for normal mammalian development and physiology. However, despite the general relevance of this process, genome stability alteration due to genetic or non-genetic conditions has a particularly profound impact on the developing cerebral cortex. In this review, we will analyze the main pathways involved in maintenance of genome stability, the consequences of their alterations with regard to central nervous system development, as well as the possible molecular and cellular basis of this specificity.
- Published
- 2018
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48. Citron kinase-dependent F-actin maintenance at midbody secondary ingression sites mediates abscission.
- Author
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Dema A, Macaluso F, Sgrò F, Berto GE, Bianchi FT, Chiotto AA, Pallavicini G, Di Cunto F, and Gai M
- Subjects
- Humans, Actins metabolism, Cytokinesis physiology, Intracellular Signaling Peptides and Proteins metabolism, Protein Serine-Threonine Kinases metabolism
- Abstract
Abscission is the final step of cytokinesis whereby the intercellular bridge (ICB) linking the two daughter cells is cut. The ICB contains a structure called the midbody, required for the recruitment and organization of the abscission machinery. Final midbody severing is mediated by formation of secondary midbody ingression sites, where the ESCRT III component CHMP4B is recruited to mediate membrane fusion. It is presently unknown how cytoskeletal elements cooperate with CHMP4B to mediate abscission. Here, we show that F-actin is associated with midbody secondary sites and is necessary for abscission. F-actin localization at secondary sites depends on the activity of RhoA and on the abscission regulator citron kinase (CITK). CITK depletion accelerates loss of F-actin proteins at the midbody and subsequent cytokinesis defects are reversed by restoring actin polymerization. Conversely, midbody hyperstabilization produced by overexpression of CITK and ANLN is reversed by actin depolymerization. CITK is required for localization of F-actin and ANLN at the abscission sites, as well as for CHMP4B recruitment. These results indicate that control of actin dynamics downstream of CITK prepares the abscission site for the final cut., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)
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- 2018
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49. IRF4 Mediates the Oncogenic Effects of STAT3 in Anaplastic Large Cell Lymphomas.
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Bandini C, Pupuleku A, Spaccarotella E, Pellegrino E, Wang R, Vitale N, Duval C, Cantarella D, Rinaldi A, Provero P, Di Cunto F, Medico E, Bertoni F, Inghirami G, and Piva R
- Abstract
Systemic anaplastic large cell lymphomas (ALCL) are a category of T-cell non-Hodgkin's lymphomas which can be divided into anaplastic lymphoma kinase (ALK) positive and ALK negative subgroups, based on ALK gene rearrangements. Among several pathways aberrantly activated in ALCL, the constitutive activation of signal transducer and activator of transcription 3 (STAT3) is shared by all ALK positive ALCL and has been detected in a subgroup of ALK negative ALCL. To discover essential mediators of STAT3 oncogenic activity that may represent feasible targets for ALCL therapies, we combined gene expression profiling analysis and RNA interference functional approaches. A shRNA screening of STAT3-modulated genes identified interferon regulatory factor 4 (IRF4) as a key driver of ALCL cell survival. Accordingly, ectopic IRF4 expression partially rescued STAT3 knock-down effects. Treatment with immunomodulatory drugs (IMiDs) induced IRF4 down regulation and resulted in cell death, a phenotype rescued by IRF4 overexpression. However, the majority of ALCL cell lines were poorly responsive to IMiDs treatment. Combination with JQ1, a bromodomain and extra-terminal (BET) family antagonist known to inhibit MYC and IRF4, increased sensitivity to IMiDs. Overall, these results show that IRF4 is involved in STAT3-oncogenic signaling and its inhibition provides alternative avenues for the design of novel/combination therapies of ALCL., Competing Interests: The authors have no conflicts of interest.
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- 2018
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50. ASPM and CITK regulate spindle orientation by affecting the dynamics of astral microtubules.
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Gai M, Bianchi FT, Vagnoni C, Vernì F, Bonaccorsi S, Pasquero S, Berto GE, Sgrò F, Chiotto AA, Annaratone L, Sapino A, Bergo A, Landsberger N, Bond J, Huttner WB, and Di Cunto F
- Published
- 2017
- Full Text
- View/download PDF
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