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1. Somatic genetic rescue of a germline ribosome assembly defect

Author

Shengjiang Tan, Laëtitia Kermasson, Christine Hilcenko, Vasileios Kargas, David Traynor, Ahmed Z. Boukerrou, Norberto Escudero-Urquijo, Alexandre Faille, Alexis Bertrand, Maxim Rossmann, Beatriz Goyenechea, Li Jin, Jonathan Moreil, Olivier Alibeu, Blandine Beaupain, Christine Bôle-Feysot, Stefano Fumagalli, Sophie Kaltenbach, Jean-Alain Martignoles, Cécile Masson, Patrick Nitschké, Mélanie Parisot, Aurore Pouliet, Isabelle Radford-Weiss, Frédéric Tores, Jean-Pierre de Villartay, Mohammed Zarhrate, Ai Ling Koh, Kong Boo Phua, Bruno Reversade, Peter J. Bond, Christine Bellanné-Chantelot, Isabelle Callebaut, François Delhommeau, Jean Donadieu, Alan J. Warren, and Patrick Revy

Subjects
Science
Abstract

Shwachman-Diamond syndrome (SDS) is a leukemia predisposition disorder that is caused by defective release of eIF6 during ribosome assembly. Here the authors show that acquired somatic EIF6 mutations are frequent in the hematopoietic cells from individuals with SDS and provide a selective advantage over non-modified cells.

Published
2021
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2. Publisher Correction: Somatic genetic rescue of a germline ribosome assembly defect

Author

Shengjiang Tan, Laëtitia Kermasson, Christine Hilcenko, Vasileios Kargas, David Traynor, Ahmed Z. Boukerrou, Norberto Escudero-Urquijo, Alexandre Faille, Alexis Bertrand, Maxim Rossmann, Beatriz Goyenechea, Li Jin, Jonathan Moreil, Olivier Alibeu, Blandine Beaupain, Christine Bôle-Feysot, Stefano Fumagalli, Sophie Kaltenbach, Jean-Alain Martignoles, Cécile Masson, Patrick Nitschké, Mélanie Parisot, Aurore Pouliet, Isabelle Radford-Weiss, Frédéric Tores, Jean-Pierre de Villartay, Mohammed Zarhrate, Ai Ling Koh, Kong Boo Phua, Bruno Reversade, Peter J. Bond, Christine Bellanné-Chantelot, Isabelle Callebaut, François Delhommeau, Jean Donadieu, Alan J. Warren, and Patrick Revy

Subjects
Science
Published
2022
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3. Role of miR-146a in neural stem cell differentiation and neural lineage determination: relevance for neurodevelopmental disorders

Author

Lam Son Nguyen, Julien Fregeac, Christine Bole-Feysot, Nicolas Cagnard, Anand Iyer, Jasper Anink, Eleonora Aronica, Olivier Alibeu, Patrick Nitschke, and Laurence Colleaux

Subjects
Autism spectrum disorders, microRNA, Human neural stem cell, Transcriptome, Neurology. Diseases of the nervous system, RC346-429
Abstract

Abstract Background MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression at the post-transcriptional level. miRNAs have emerged as important modulators of brain development and neuronal function and are implicated in several neurological diseases. Previous studies found miR-146a upregulation is the most common miRNA deregulation event in neurodevelopmental disorders such as autism spectrum disorder (ASD), epilepsy, and intellectual disability (ID). Yet, how miR-146a upregulation affects the developing fetal brain remains unclear. Methods We analyzed the expression of miR-146a in the temporal lobe of ASD children using Taqman assay. To assess the role of miR-146a in early brain development, we generated and characterized stably induced H9 human neural stem cell (H9 hNSC) overexpressing miR-146a using various cell and molecular biology techniques. Results We first showed that miR-146a upregulation occurs early during childhood in the ASD brain. In H9 hNSC, miR-146a overexpression enhances neurite outgrowth and branching and favors differentiation into neuronal like cells. Expression analyses revealed that 10% of the transcriptome was deregulated and organized into two modules critical for cell cycle control and neuronal differentiation. Twenty known or predicted targets of miR-146a were significantly deregulated in the modules, acting as potential drivers. The two modules also display distinct transcription profiles during human brain development, affecting regions relevant for ASD including the neocortex, amygdala, and hippocampus. Cell type analyses indicate markers for pyramidal, and interneurons are highly enriched in the deregulated gene list. Up to 40% of known markers of newly defined neuronal lineages were deregulated, suggesting that miR-146a could participate also in the acquisition of neuronal identities. Conclusion Our results demonstrate the dynamic roles of miR-146a in early neuronal development and provide new insight into the molecular events that link miR-146a overexpression to impaired neurodevelopment. This, in turn, may yield new therapeutic targets and strategies.

Published
2018
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4. Somatic genetic rescue of a germline ribosome assembly defect

Author

Vasileios Kargas, Mélanie Parisot, Norberto Escudero-Urquijo, Alexis Bertrand, Christine Bellanné-Chantelot, Jean Donadieu, Mohammed Zarhrate, Patrick Nitschke, Cécile Masson, Laëtitia Kermasson, Beatriz Goyenechea, Sophie Kaltenbach, Alan J. Warren, David Traynor, Stefano Fumagalli, Li Jin, Blandine Beaupain, Bruno Reversade, Ahmed Z. Boukerrou, Peter J. Bond, M. Rossmann, Olivier Alibeu, Jean-Alain Martignoles, Christine Bole-Feysot, Jonathan Moreil, Shengjiang Tan, François Delhommeau, Patrick Revy, Kong Boo Phua, Alexandre Faille, Aurore Pouliet, Christine Hilcenko, Frédéric Tores, Isabelle Radford-Weiss, Ai Ling Koh, Isabelle Callebaut, Jean-Pierre de Villartay, Cambridge Institute for Medical Research (CIMR), University of Cambridge [UK] (CAM), Wellcome Trust-Medical Research Council Cambridge Stem Cell Institute, Imagine - Institut des maladies génétiques (IHU) (Imagine - U1163), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), MRC Laboratory of Molecular Biology [Cambridge, UK] (LMB), University of Cambridge [UK] (CAM)-Medical Research Council, Structure Fédérative de Recherche Necker (SFR Necker - UMS 3633 / US24), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Institut Necker Enfants-Malades (INEM - UM 111 (UMR 8253 / U1151)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Université Paris Descartes - Paris 5 (UPD5), CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Centre de Recherche Saint-Antoine (CR Saint-Antoine), Sorbonne Université (SU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), KK Women's and Children's Hospital [Singapore], Genome Institute of Singapore (GIS), Bioinformatics Institute [Singapore], Agency for science, technology and research [Singapore] (A*STAR), National University of Singapore (NUS), CHU Pitié-Salpêtrière [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Sorbonne Université (SU), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Institut de recherche pour le développement [IRD] : UR206-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Babraham Research Campus [Cambridge, Royaume-Uni], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Université Sorbonne Paris Cité (USPC), Centre de Recherche Saint-Antoine (CRSA), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), ACS - Heart failure & arrhythmias, ARD - Amsterdam Reproduction and Development, Escudero-Urquijo, Norberto [0000-0002-8201-5884], Parisot, Mélanie [0000-0003-4312-2035], Reversade, Bruno [0000-0002-4070-7997], Bond, Peter J. [0000-0003-2900-098X], Bellanné-Chantelot, Christine [0000-0001-8415-6771], Warren, Alan J. [0000-0001-9277-4553], Revy, Patrick [0000-0003-0758-8022], Apollo - University of Cambridge Repository, Service de Génétique Cytogénétique et Embryologie [CHU Pitié-Salpêtrière], Hilcenko, Christine [0000-0002-9596-7833], Rossmann, Maxim [0000-0001-8811-3277], Bond, Peter J [0000-0003-2900-098X], Warren, Alan [0000-0001-9277-4553], Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPC), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPC), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPC), HAL-SU, Gestionnaire, ANR-18-IDEX-0001,Université de Paris,Université de Paris(2018), Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), and Warren, Alan J [0000-0001-9277-4553]

Subjects
Somatic cell, [SDV]Life Sciences [q-bio], General Physics and Astronomy, [SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC], Germline, 631/208/737, 692/699/1541, Ribosome assembly, 13/1, 0302 clinical medicine, hemic and lymphatic diseases, 38/22, Dictyostelium, Eukaryotic Initiation Factors, Child, Cells, Cultured, Biological Phenomena, Genetics, 0303 health sciences, Multidisciplinary, 631/208/514/2254, article, [SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology, Ribosome Subunits, Large, Eukaryotic, Ribosome, Shwachman-Diamond Syndrome, 3. Good health, [SDV] Life Sciences [q-bio], 64/24, [SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology, 030220 oncology & carcinogenesis, Child, Preschool, Drosophila, Haematological diseases, Protein Binding, Adult, congenital, hereditary, and neonatal diseases and abnormalities, Adolescent, Science, 45/23, Saccharomyces cerevisiae, Biology, Molecular Dynamics Simulation, General Biochemistry, Genetics and Molecular Biology, 82/80, 03 medical and health sciences, Young Adult, Germline mutation, Animals, Humans, Gene, Ribonucleoprotein, U5 Small Nuclear, 030304 developmental biology, Sequence Homology, Amino Acid, Eukaryotic Large Ribosomal Subunit, Point mutation, Infant, Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Chemistry, SBDS, 96/44, Peptide Elongation Factors, Germ Cells, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, Protein Biosynthesis, Mutation, Next-generation sequencing, 631/337/574/1789, Ribosomes
Abstract

Indirect somatic genetic rescue (SGR) of a germline mutation is thought to be rare in inherited Mendelian disorders. Here, we establish that acquired mutations in the EIF6 gene are a frequent mechanism of SGR in Shwachman-Diamond syndrome (SDS), a leukemia predisposition disorder caused by a germline defect in ribosome assembly. Biallelic mutations in the SBDS or EFL1 genes in SDS impair release of the anti-association factor eIF6 from the 60S ribosomal subunit, a key step in the translational activation of ribosomes. Here, we identify diverse mosaic somatic genetic events (point mutations, interstitial deletion, reciprocal chromosomal translocation) in SDS hematopoietic cells that reduce eIF6 expression or disrupt its interaction with the 60S subunit, thereby conferring a selective advantage over non-modified cells. SDS-related somatic EIF6 missense mutations that reduce eIF6 dosage or eIF6 binding to the 60S subunit suppress the defects in ribosome assembly and protein synthesis across multiple SBDS-deficient species including yeast, Dictyostelium and Drosophila. Our data suggest that SGR is a universal phenomenon that may influence the clinical evolution of diverse Mendelian disorders and support eIF6 suppressor mimics as a therapeutic strategy in SDS., Shwachman-Diamond syndrome (SDS) is a leukemia predisposition disorder that is caused by defective release of eIF6 during ribosome assembly. Here the authors show that acquired somatic EIF6 mutations are frequent in the hematopoietic cells from individuals with SDS and provide a selective advantage over non-modified cells.

Published
2021

5. Optimization of CRISPR/Cas9 Delivery to Human Hematopoietic Stem and Progenitor Cells for Therapeutic Genomic Rearrangements

Author

Cécile Masson, Mario Amendola, Annarita Miccio, Annalisa Lattanzi, Ciaran M. Lee, Chiara Antoniani, Sophie Ramadier, Gang Bao, Olivier Alibeu, Vasco Meneghini, Tristan Felix, Matthew H. Porteus, Giulia Pavani, Fulvio Mavilio, Fatima Amor, Approches génétiques intégrées et nouvelles thérapies pour les maladies rares (INTEGRARE), Université d'Évry-Val-d'Essonne (UEVE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Généthon-École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Généthon, Chromatin and gene regulation during development (Equipe Inserm U1163), Imagine - Institut des maladies génétiques (IHU) (Imagine - U1163), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Plateforme Bioinformatique [SFR Necker] (BIP-D), Structure Fédérative de Recherche Necker (SFR Necker - UMS 3633 / US24), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Rice University [Houston], Stanford University, Università degli Studi di Modena e Reggio Emilia (UNIMORE), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université d'Évry-Val-d'Essonne (UEVE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Généthon, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Università degli Studi di Modena e Reggio Emilia = University of Modena and Reggio Emilia (UNIMORE), CCSD, Accord Elsevier, École Pratique des Hautes Études (EPHE), Chromatin and gene regulation during development, and ANR-16-CE18-0004,GETH,L'Édition Génomique comme outil Thérapeutique contre les ß-Hémoglobinopathies(2016)

Subjects
[SDV]Life Sciences [q-bio], Genetic enhancement, Anemia, Sickle Cell, Biology, Heterogeneous-Nuclear Ribonucleoproteins, Viral vector, 03 medical and health sciences, 0302 clinical medicine, Genome editing, CRISPR-Associated Protein 9, hemic and lymphatic diseases, Drug Discovery, Genetics, Humans, CRISPR, Progenitor cell, Molecular Biology, 030304 developmental biology, Gene Editing, Pharmacology, 0303 health sciences, Cas9, beta-Thalassemia, Genetic Therapy, Hematopoietic Stem Cells, 3. Good health, Cell biology, Hemoglobinopathies, [SDV] Life Sciences [q-bio], Haematopoiesis, 030220 oncology & carcinogenesis, Molecular Medicine, Original Article, CRISPR-Cas Systems, Stem cell, Plasmids, RNA, Guide, Kinetoplastida, CRISPR/Cas9 delivery, genome editing, β-hemoglobinopathies, Drug Discovery3003 Pharmaceutical Science
Abstract

International audience; Editing the beta-globin locus in hematopoietic stem cells is an alternative therapeutic approach for gene therapy of beta-thalassemia and sickle cell disease. Using the CRISPR/Cas9 system, we genetically modified human hematopoietic stem and progenitor cells (HSPCs) to mimic the large rearrangements in the beta-globin locus associated with hereditary persistence of fetal hemoglobin (HPFH), a condition that mitigates the clinical phenotype of patients with beta-hemoglobinopathies. We optimized and compared the efficiency of plasmid-, lentiviral vector (LV)-, RNA-, and ribonucleoprotein complex (RNP)-based methods to deliver the CRISPR/Cas9 system into HSPCs. Plasmid delivery of Cas9 and gRNA pairs targeting two HPFH-like regions led to high frequency of genomic rearrangements and HbF reactivation in erythroblasts derived from sorted, Cas9(+) HSPCs but was associated with significant cell toxicity. RNA-mediated delivery of CRISPR/Cas9 was similarly toxic but much less efficient in editing the beta-globin locus. Transduction of HSPCs by LVs expressing Cas9 and gRNA pairs was robust and minimally toxic but resulted in poor genome-editing efficiency. Ribonucleoprotein (RNP)-based delivery of CRISPR/Cas9 exhibited a good balance between cytotoxicity and efficiency of genomic rearrangements as compared to the other delivery systems and resulted in HbF upregulation in erythroblasts derived from unselected edited HSPCs.

Published
2019

6. Corrigendum to: Diagnostic Yield of Next-Generation Sequencing in Very Early-Onset Inflammatory Bowel Diseases: A Multicenter Study

Author

Olivier Alibeu, P. Tounian, Nadia Siala, Michela Tempia-Caliera, Jean-Pierre Hugot, Sabine Rakotobe, Christelle Lenoir, Anne Breton, Caterina Strisciuglio, Víctor Manuel Navas-López, Jan Melek, Alain Fischer, Frédéric Rieux-Laucat, Marie-Claude Stolzenberg, Eric Jeziorski, Yago González-Lama, Bénédicte Pigneur, Mongi Ben Hariz, Marina Aloi, Sylvain Latour, Fabienne Mazerolles, Christian Breuer, Julie Bruneau, Clara Crémilleux, Cecile Pelatan, Vaidotas Urbonas, Alexandre Fabre, Nadine Cerf-Bensussan, Frank M. Ruemmele, Luisa Mearin, Capucine Picard, Georgia Malamut, Neslihan Gurcan, Anders Paerregaard, Isabel Pinto Pais, Dan Turner, István Máttyus, Julie Rebeuh, Jiri Bronsky, Sylvain Hanein, Peter Lewindon, Rémi Duclaux-Loras, Graziella Guariso, Anne Bourrier, Odul Egritas Gurkan, Janos Major, Stéphanie Willot, Mara Cananzi, Marianna Parlato, Claudio Romano, Alain Lachaux, Matjaz Homan, Jorge Amil Dias, Eva Lévy, A Fischer, Stéphanie Coopman, Jan Krzysztof Nowak, Fernando Magro, Clémentine Dumant-Forest, Stephan Buderus, Bernadette Bègue, Fabienne Charbit-Henrion, Olivier Goulet, Evi Karanika, Alain Dabadie, Emmanuel Mas, Marta German Diaz, Cécile Fourrage, Rosa Lima, Charbit-Henrion, Fabienne, Parlato, Marianna, Hanein, Sylvain, Duclaux-Loras, Rémi, Nowak, Jan, Begue, Bernadette, Rakotobe, Sabine, Bruneau, Julie, Fourrage, Cécile, Alibeu, Olivier, Rieux-Laucat, Frédéric, Lévy, Eva, Stolzenberg, Marie-Claude, Mazerolles, Fabienne, Latour, Sylvain, Lenoir, Christelle, Fischer, Alain, Picard, Capucine, Aloi, Marina, Dias, Jorge Amil, Hariz, Mongi Ben, Bourrier, Anne, Breuer, Christian, Breton, Anne, Bronsky, Jiri, Buderus, Stephan, Cananzi, Mara, Coopman, Stéphanie, Crémilleux, Clara, Dabadie, Alain, Dumant-Forest, Clémentine, Gurkan, Odul Egrita, Fabre, Alexandre, Fischer, Aude, Diaz, Marta German, Gonzalez-Lama, Yago, Goulet, Olivier, Guariso, Graziella, Gurcan, Neslihan, Homan, Matjaz, Hugot, Jean-Pierre, Jeziorski, Eric, Karanika, Evi, Lachaux, Alain, Lewindon, Peter, Lima, Rosa, Magro, Fernando, Major, Jano, Malamut, Georgia, Mas, Emmanuel, Mattyus, Istvan, Mearin, Luisa M, Melek, Jan, Navas-Lopez, Victor Manuel, Paerregaard, Ander, Pelatan, Cecile, Pigneur, Bénédicte, Pais, Isabel Pinto, Rebeuh, Julie, Romano, Claudio, Siala, Nadia, Strisciuglio, Caterina, Tempia-Caliera, Michela, Tounian, Patrick, Turner, Dan, Urbonas, Vaidota, Willot, Stéphanie, Ruemmele, Frank M, and Cerf-Bensussan, Nadine

Subjects
VEO-IBD, business.industry, Yield (finance), monogenic IBD, next generation sequencing, very early onset IBD, Gastroenterology, Inflammatory Bowel Diseases, Genetics and molecular epidemiology, Original Articles, General Medicine, monogenic disorders, Bioinformatics, Very early onset, DNA sequencing, paediatrics, Editor's Choice, Multicenter study, TNGS, Medicine, genetics and molecular epidemiology, business
Abstract

Background and Aims An expanding number of monogenic defects have been identified as causative of severe forms of very early-onset inflammatory bowel diseases [VEO-IBD]. The present study aimed at defining how next-generation sequencing [NGS] methods can be used to improve identification of known molecular diagnosis and to adapt treatment. Methods A total of 207 children were recruited in 45 paediatric centres through an international collaborative network [ESPGHAN GENIUS working group] with a clinical presentation of severe VEO-IBD [n = 185] or an anamnesis suggestive of a monogenic disorder [n = 22]. Patients were divided at inclusion into three phenotypic subsets: predominantly small bowel inflammation, colitis with perianal lesions, and colitis only. Methods to obtain molecular diagnosis included functional tests followed by specific Sanger sequencing, custom-made targeted NGS, and in selected cases whole exome sequencing [WES] of parents-child trios. Genetic findings were validated clinically and/or functionally. Results Molecular diagnosis was achieved in 66/207 children [32%]: 61% with small bowel inflammation, 39% with colitis and perianal lesions, and 18% with colitis only. Targeted NGS pinpointed gene mutations causative of atypical presentations, and identified large exonic copy number variations previously missed by WES. Conclusions Our results lead us to propose an optimised diagnostic strategy to identify known monogenic causes of severe IBD.

Published
2020

7. Targeted Exome Sequencing Identifies PBX1 as Involved in Monogenic Congenital Anomalies of the Kidney and Urinary Tract

Author

Olivier Alibeu, Cécile Jeanpierre, Marc Jeanpierre, Cécile Fourrage, Valérie Malan, Charline Henry, Robert Novo, Frédéric Tores, Vincent Morinière, Madeline Louise Reilly, Marc Bras, Sophie Saunier, Rémi Salomon, Corinne Antignac, Camille Humbert, Lara De Tomasi, Patrick Nitschke, Dominique Gaillard, Laurence Heidet, Marie Gonzales, Christine Pietrement, Jelena Martinovic, Joelle Roume, Christine Bole-Feysot, and Elise Schaefer

Subjects
0301 basic medicine, Genetics, Candidate gene, Genetic heterogeneity, Kidney development, General Medicine, Disease, Biology, 03 medical and health sciences, 030104 developmental biology, Nephrology, Cancer research, Epigenetics, Gene, Loss function, Exome sequencing
Abstract

Congenital anomalies of the kidney and urinary tract (CAKUT) occur in three to six of 1000 live births, represent about 20% of the prenatally detected anomalies, and constitute the main cause of CKD in children. These disorders are phenotypically and genetically heterogeneous. Monogenic causes of CAKUT in humans and mice have been identified. However, despite high-throughput sequencing studies, the cause of the disease remains unknown in most patients, and several studies support more complex inheritance and the role of environmental factors and/or epigenetics in the pathophysiology of CAKUT. Here, we report the targeted exome sequencing of 330 genes, including genes known to be involved in CAKUT and candidate genes, in a cohort of 204 unrelated patients with CAKUT; 45% of the patients were severe fetal cases. We identified pathogenic mutations in 36 of 204 (17.6%) patients. These mutations included five de novo heterozygous loss of function mutations/deletions in the PBX homeobox 1 gene (PBX1), a gene known to have a crucial role in kidney development. In contrast, the frequency of SOX17 and DSTYK variants recently reported as pathogenic in CAKUT did not indicate causality. These findings suggest that PBX1 is involved in monogenic CAKUT in humans and call into question the role of some gene variants recently reported as pathogenic in CAKUT. Targeted exome sequencing also proved to be an efficient and cost-effective strategy to identify pathogenic mutations and deletions in known CAKUT genes.

Published
2017

8. Early acute microvascular kidney transplant rejection in the absence of anti-HLA antibodies is associated with preformed IgG antibodies against diverse glomerular endothelial cell antigens

Author

Olivier Thaunat, Nicolas Cagnard, Cyril Garrouste, Marc Hazzan, Christophe Mariat, Marion Rabant, Nadia Arzouk, Pierre Merville, Emmanuel Zorn, Jean Luc Taupin, Pierre François Westeel, Jean-Paul Duong Van Huyen, Carole Burger, Alain Le Moine, Philippe Gatault, Nassim Kamar, Olivier Alibeu, Dominique Bertrand, Simon C. Satchell, Sarah S.B. See, Sophie Caillard, Joseph Rivalan, Baptiste Lamarthée, Christophe Legendre, Marc Ladrière, Béatrice Charreau, Vincent Vuiblet, Magali Giral, Marie Matignon, Dany Anglicheau, Johnny Sayegh, Alexandre Hertig, Anne Cesbron, Sylvain Pagie, and Marianne Delville

Subjects
Adult, Graft Rejection, Male, Vasculitis, Thrombotic microangiopathy, Time Factors, Cell, Kidney Glomerulus, Bristol Heart Institute, 030232 urology & nephrology, Hemorrhage, 030230 surgery, Receptor, Angiotensin, Type 1, 03 medical and health sciences, 0302 clinical medicine, Antigen, Clinical Research, Biopsy, Medicine, Humans, Receptor, Aged, biology, medicine.diagnostic_test, Endothelin-1, business.industry, Thrombotic Microangiopathies, Endothelial Cells, General Medicine, Middle Aged, medicine.disease, Kidney Transplantation, Transplantation, medicine.anatomical_structure, Nephrology, Immunoglobulin G, Immunology, Acute Disease, Microvessels, biology.protein, Female, Antibody, business
Abstract

BACKGROUND: Although anti-HLA antibodies (Abs) cause most antibody-mediated rejections of renal allografts, non-anti-HLA Abs have also been postulated to contribute. A better understanding of such Abs in rejection is needed.METHODS: We conducted a nationwide study to identify kidney transplant recipients without anti-HLA donor-specific Abs who experienced acute graft dysfunction within 3 months after transplantation and showed evidence of microvascular injury, called acute microvascular rejection (AMVR). We developed a crossmatch assay to assess serum reactivity to human microvascular endothelial cells, and used a combination of transcriptomic and proteomic approaches to identify non-HLA Abs.RESULTS: We identified a highly selected cohort of 38 patients with early acute AMVR. Biopsy specimens revealed intense microvascular inflammation and the presence of vasculitis (in 60.5%), interstitial hemorrhages (31.6%), or thrombotic microangiopathy (15.8%). Serum samples collected at the time of transplant showed that previously proposed anti-endothelial cell Abs-angiotensin type 1 receptor (AT1R), endothelin-1 type A and natural polyreactive Abs-did not increase significantly among patients with AMVR compared with a control group of stable kidney transplant recipients. However, 26% of the tested AMVR samples were positive for AT1R Abs when a threshold of 10 IU/ml was used. The crossmatch assay identified a common IgG response that was specifically directed against constitutively expressed antigens of microvascular glomerular cells in patients with AMVR. Transcriptomic and proteomic analyses identified new targets of non-HLA Abs, with little redundancy among individuals.CONCLUSIONS: Our findings indicate that preformed IgG Abs targeting non-HLA antigens expressed on glomerular endothelial cells are associated with early AMVR, and that in vitro cell-based assays are needed to improve risk assessments before transplant.

Published
2019

9. Loss of ARHGEF1 causes a human primary antibody deficiency

Author

Anne Durandy, Sébastien Lofek, Hicham Lamrini, Eric Oksenhendler, Loic Chentout, Isabelle André-Schmutz, Elizabeth Macintyre, Sven Kracker, Marie-Céline Deau, Marc Bras, Lucie Heurtier, Amélie Trinquand, Marina Cavazzana, Amine Bouafia, Véronique Meignin, Olivier Alibeu, Alain Fischer, Julie Bruneau, Thierry Jo Molina, Capucine Picard, Institut Cochin (IC UM3 (UMR 8104 / U1016)), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), CHU Necker - Enfants Malades [AP-HP], Institut Necker Enfants-Malades (INEM - UM 111 (UMR 8253 / U1151)), Laboratoire d'Anatomie Pathologique, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Génétique Humaine des Maladies Infectieuses (Inserm U980), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5), Service hématologie, Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Necker - Enfants Malades [AP-HP], Imagine - Institut des maladies génétiques (IMAGINE - U1163), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Plateforme de bioinformatique (UNIV Paris Descartes), Université Paris Descartes - Paris 5 (UPD5), Hôpital Hôtel Dieu, Département de Biothérapie [CHU Necker], CHU Necker - Enfants Malades [AP-HP]-Université Paris Descartes - Paris 5 (UPD5)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Developpement Normal et Pathologique du Système Immunitaire, Service d'Immunopathologie Clinique, ANR-15-CE15-0020,PIKimun,Caractérisation de la voie PI3K / AKT / mTOR - dans les lymphocytes humains(2015), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5), Chaire Médecine expérimentale (A. Fischer), and Collège de France (CdF (institution))

Subjects
0301 basic medicine, Male, RHOA, Primary Immunodeficiency Diseases, T-Lymphocytes, [SDV]Life Sciences [q-bio], GTP-Binding Protein alpha Subunits, G12-G13, 03 medical and health sciences, Phosphatidylinositol 3-Kinases, 0302 clinical medicine, medicine, Humans, Lymphocytes, Protein kinase B, PI3K/AKT/mTOR pathway, B cell, ComputingMilieux_MISCELLANEOUS, B-Lymphocytes, rho-Associated Kinases, [SDV.GEN]Life Sciences [q-bio]/Genetics, biology, Chemistry, Siblings, Germinal center, General Medicine, Marginal zone, Actin cytoskeleton, Germinal Center, Molecular biology, 3. Good health, Haematopoiesis, 030104 developmental biology, medicine.anatomical_structure, [SDV.IMM.IA]Life Sciences [q-bio]/Immunology/Adaptive immunology, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, 030220 oncology & carcinogenesis, biology.protein, Commentary, [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, rhoA GTP-Binding Protein, Immunologic Memory, Proto-Oncogene Proteins c-akt, Rho Guanine Nucleotide Exchange Factors, Research Article, Signal Transduction
Abstract

ARHGEF1 is a RhoA-specific guanine nucleotide exchange factor expressed in hematopoietic cells. We used whole-exome sequencing to identify compound heterozygous mutations in ARHGEF1, resulting in the loss of ARHGEF1 protein expression in 2 primary antibody–deficient siblings presenting with recurrent severe respiratory tract infections and bronchiectasis. Both ARHGEF1-deficient patients showed an abnormal B cell immunophenotype, with a deficiency in marginal zone and memory B cells and an increased frequency of transitional B cells. Furthermore, the patients’ blood contained immature myeloid cells. Analysis of a mediastinal lymph node from one patient highlighted the small size of the germinal centers and an abnormally high plasma cell content. On the molecular level, T and B lymphocytes from both patients displayed low RhoA activity and low steady-state actin polymerization (even after stimulation of lysophospholipid receptors). As a consequence of disturbed regulation of the RhoA downstream target Rho-associated kinase I/II (ROCK), the patients’ lymphocytes failed to efficiently restrain AKT phosphorylation. Enforced ARHGEF1 expression or drug-induced activation of RhoA in the patients’ cells corrected the impaired actin polymerization and AKT regulation. Our results indicate that ARHGEF1 activity in human lymphocytes is involved in controlling actin cytoskeleton dynamics, restraining PI3K/AKT signaling, and confining B lymphocytes and myelocytes within their dedicated functional environment.

Published
2019

10. Heterozygous Mutations in MAP3K7 , Encoding TGF-β-Activated Kinase 1, Cause Cardiospondylocarpofacial Syndrome

Author

Capucine Picard, Carine Le Goff, Graziella Pinto, Olivier Alibeu, Arnold Munnich, Patrick Nistchke, Wilfried Le Goff, Damien Bonnet, Valérie Cormier-Daire, Maya Chrabieh, and Curtis Rogers

Subjects
Male, 0301 basic medicine, Hearing Loss, Conductive, Interleukin-1beta, 030105 genetics & heredity, medicine.disease_cause, p38 Mitogen-Activated Protein Kinases, Transforming Growth Factor beta, Genetics(clinical), Child, Genetics (clinical), Mutation, Kinase, Mitral Valve Insufficiency, Syndrome, MAP Kinase Kinase Kinases, Child, Preschool, Cervical Vertebrae, Female, Osteosclerosis, Adult, Heterozygote, medicine.medical_specialty, Adolescent, MAP Kinase Signaling System, Biology, MAP3K7, Hearing Loss, Bilateral, Young Adult, 03 medical and health sciences, Report, Internal medicine, Genetics, medicine, Humans, Abnormalities, Multiple, Protein kinase A, Carpal Bones, MAP kinase kinase kinase, Brachydactyly, Tarsal Bones, Transforming growth factor beta, Fibroblasts, medicine.disease, 030104 developmental biology, Endocrinology, Gene Expression Regulation, Cancer research, biology.protein, Transforming growth factor
Abstract

Cardiospondylocarpofacial (CSCF) syndrome is characterized by growth retardation, dysmorphic facial features, brachydactyly with carpal-tarsal fusion and extensive posterior cervical vertebral synostosis, cardiac septal defects with valve dysplasia, and deafness with inner ear malformations. Whole-exome sequencing identified heterozygous MAP3K7 mutations in six distinct CSCF-affected individuals from four families and ranging in age from 5 to 37 years. MAP3K7 encodes transforming growth factor β (TGF-β)-activated kinase 1 (TAK1), which is involved in the mitogen-activated protein kinase (MAPK)-p38 signaling pathway. MAPK-p38 signaling was markedly altered when expression of non-canonical TGF-β-driven target genes was impaired. These findings support the loss of transcriptional control of the TGF-β-MAPK-p38 pathway in fibroblasts obtained from affected individuals. Surprisingly, although TAK1 is located at the crossroad of inflammation, immunity, and cancer, this study reports MAP3K7 mutations in a developmental disorder affecting mainly cartilage, bone, and heart.

Published
2016

11. Role of miR-146a in neural stem cell differentiation and neural lineage determination: relevance for neurodevelopmental disorders

Author

Olivier Alibeu, Lam Son Nguyen, Julien Fregeac, Eleonora Aronica, Christine Bole-Feysot, Anand Iyer, Patrick Nitschké, Laurence Colleaux, Jasper J. Anink, Nicolas Cagnard, Pathology, ANS - Cellular & Molecular Mechanisms, APH - Aging & Later Life, APH - Mental Health, Imagine - Institut des maladies génétiques (IMAGINE - U1163), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Necker - Enfants Malades [AP-HP], Université Paris Descartes - Paris 5 (UPD5), Université Sorbonne Paris Cité (USPC), Genome Analysis, Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)

Subjects
0301 basic medicine, MESH: Neural Stem Cells, Male, Human neural stem cell, Autism Spectrum Disorder, lcsh:RC346-429, Transcriptome, Neural Stem Cells, MESH: Child, MESH: Up-Regulation, Child, MESH: Autism Spectrum Disorder, Neocortex, microRNA, Human brain, Autism spectrum disorders, Neural stem cell, Temporal Lobe, Up-Regulation, Psychiatry and Mental health, medicine.anatomical_structure, Female, Cell type, Neurite, Neurogenesis, Biology, Cell Line, 03 medical and health sciences, Developmental Neuroscience, Downregulation and upregulation, medicine, MESH: Temporal Lobe, Humans, Cell Lineage, Molecular Biology, lcsh:Neurology. Diseases of the nervous system, MESH: Humans, Research, MESH: Cell Lineage, MESH: Male, MESH: Cell Line, MESH: Neurogenesis, MicroRNAs, 030104 developmental biology, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, Neuroscience, MESH: Female, MESH: MicroRNAs, Developmental Biology
Abstract

Background MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression at the post-transcriptional level. miRNAs have emerged as important modulators of brain development and neuronal function and are implicated in several neurological diseases. Previous studies found miR-146a upregulation is the most common miRNA deregulation event in neurodevelopmental disorders such as autism spectrum disorder (ASD), epilepsy, and intellectual disability (ID). Yet, how miR-146a upregulation affects the developing fetal brain remains unclear. Methods We analyzed the expression of miR-146a in the temporal lobe of ASD children using Taqman assay. To assess the role of miR-146a in early brain development, we generated and characterized stably induced H9 human neural stem cell (H9 hNSC) overexpressing miR-146a using various cell and molecular biology techniques. Results We first showed that miR-146a upregulation occurs early during childhood in the ASD brain. In H9 hNSC, miR-146a overexpression enhances neurite outgrowth and branching and favors differentiation into neuronal like cells. Expression analyses revealed that 10% of the transcriptome was deregulated and organized into two modules critical for cell cycle control and neuronal differentiation. Twenty known or predicted targets of miR-146a were significantly deregulated in the modules, acting as potential drivers. The two modules also display distinct transcription profiles during human brain development, affecting regions relevant for ASD including the neocortex, amygdala, and hippocampus. Cell type analyses indicate markers for pyramidal, and interneurons are highly enriched in the deregulated gene list. Up to 40% of known markers of newly defined neuronal lineages were deregulated, suggesting that miR-146a could participate also in the acquisition of neuronal identities. Conclusion Our results demonstrate the dynamic roles of miR-146a in early neuronal development and provide new insight into the molecular events that link miR-146a overexpression to impaired neurodevelopment. This, in turn, may yield new therapeutic targets and strategies. Electronic supplementary material The online version of this article (10.1186/s13229-018-0219-3) contains supplementary material, which is available to authorized users.

Published
2018

12. APOL1 risk genotype in European steroid-resistant nephrotic syndrome and/or focal segmental glomerulosclerosis patients of different African ancestries

Author

Olivier Alibeu, Marie Josèphe Tête, Olivier Gribouval, Olivia Boyer, Aude Servais, Alexandre Karras, Julien Hogan, Corinne Antignac, Cécile Fourrage, Bertrand Knebelmann, Jacques Dantal, and Claire Dossier

Subjects
Adult, Male, medicine.medical_specialty, Nephrotic Syndrome, Adolescent, Genotype, Apolipoprotein L1, 030232 urology & nephrology, Drug Resistance, Black People, 030204 cardiovascular system & hematology, Compound heterozygosity, Gastroenterology, Cohort Studies, 03 medical and health sciences, Young Adult, 0302 clinical medicine, Focal segmental glomerulosclerosis, Risk Factors, Internal medicine, Medicine, Humans, Minimal change disease, Genetic Predisposition to Disease, Child, Transplantation, biology, business.industry, Glomerulosclerosis, Focal Segmental, Homozygote, medicine.disease, Prognosis, Steroid-resistant nephrotic syndrome, Pedigree, Survival Rate, Nephrology, Case-Control Studies, Cohort, Mutation, biology.protein, Female, Steroids, France, business, Nephrotic syndrome
Abstract

Background Apolipoprotein L1 (APOL1) risk variants are strongly associated with sporadic focal segmental glomerulosclerosis (FSGS) in populations with African ancestry. We determined the frequency of G1/G2 variants in steroid-resistant nephrotic syndrome (SRNS)/FSGS patients with African or French West Indies ancestry in France and its relationships with other SRNS genes. Methods In a cohort of 152 patients (139 families), the APOL1 risk variants were genotyped by direct Sanger sequencing and pathogenic mutations were screened by next-generation sequencing with a panel including 35 SRNS genes. Results The two risk allele [high-risk (HR)] genotypes were found in 43.1% (66/152) of subjects compared with 18.9% (106/562) in a control population (P Conclusions The HR genotype is frequent in FSGS patients with African ancestry in our cohort, especially in those originating from the West Indies, and confer a poor renal prognosis. It is usually not associated with other causative mutations in monogenic SRNS genes.

Published
2018

13. Recessive loss of function PIGN alleles, including an intragenic deletion with founder effect in La Réunion Island, in patients with Fryns syndrome

Author

Asma Chebil, Duksha Ramful, Thibaud Jouan, Hanitra Randrianaivo, Norbert F. Ajeawung, Nicolas Gruchy, Yannis Duffourd, Bérénice Doray, Christopher T. Gordon, Francesca Filippini, Christine Bole-Feysot, Philippe M. Campeau, Asma Omarjee, Julien Thevenon, Paul Kuentz, Jean-Luc Alessandri, Patrick Nitschke, Laurence Faivre, Jeanne Amiel, Coralie Dumont, Thi Tuyet Mai Nguyen, Marion Gérard, Myriam Oufadem, Guillaume Benoist, Olivier Alibeu, Marie-Line Jacquemont, Centre Hospitalier Universitaire de La Réunion (CHU La Réunion), Imagine - Institut des maladies génétiques (IMAGINE - U1163), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Sud Saint Pierre [Ile de la Réunion], Service de Génétique [CHU Caen], Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Tumorothèque de Caen Basse-Normandie (TCBN), Centre de recherche du CHU Sainte-Justine, CHU Sainte Justine [Montréal], Centre Hospitalier de Mayotte, Equipe GAD (LNC - U1231), Lipides - Nutrition - Cancer [Dijon - U1231] (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, FHU TRANSLAD, Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Laboratoire de Génétique Médicale (LGM), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche du CHU Sainte-Justine [Montreal], Université de Montréal (UdeM)-CHU Sainte Justine [Montréal], Service de réanimation néonatologique et pédiatrique [La Réunion], Groupe Hospitalier Sud-CHR La réunion, Embryology and genetics of human malformation (Equipe Inserm U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Génétique et épigénétique des maladies métaboliques, neurosensorielles et du développement (Inserm U781), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Université de Caen Normandie (UNICAEN), Normandie Université (NU), Biologie, génétique et thérapies ostéoarticulaires et respiratoires (BIOTARGEN), Normandie Université (NU)-Normandie Université (NU), Pôle Gynécologie Obstétrique [CHM, Mayotte], Centre de génétique - Centre de référence des maladies rares, anomalies du développement et syndromes malformatifs (CHU de Dijon), Hôpital d'Enfants [CHU Dijon], Hôpital du Bocage, Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon)-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Pôle Femme-Mère-Enfant [CHU La Réunion, Saint-Pierre, La Réunion], Plateforme de génomique [SFR Necker], Structure Fédérative de Recherche Necker (SFR Necker - UMS 3633 / US24), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Plate Forme Paris Descartes de Bioinformatique (BIP-D), Université Paris Descartes - Paris 5 (UPD5), CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Biologie moléculaire et cellulaire de la différenciation, Université Joseph Fourier - Grenoble 1 (UJF)-Institut Albert Bonniot-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Universitaire de La Réunion ( CHU La Réunion ), Imagine - Institut des maladies génétiques ( IMAGINE - U1163 ), Centre National de la Recherche Scientifique ( CNRS ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université Paris Descartes - Paris 5 ( UPD5 ), Service de Génétique Clinique [Caen], Université de Caen Normandie ( UNICAEN ), Normandie Université ( NU ) -Normandie Université ( NU ) -CHU Caen, Lipides - Nutrition - Cancer [Dijon - U1231] ( LNC ), Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Institut National de la Santé et de la Recherche Médicale ( INSERM ), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Laboratoire de Génétique Médicale ( LGM ), Université de Strasbourg ( UNISTRA ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ), Service de Réanimation Néonatale et Pédiatrique, CHD Félix Guyon, Eidgenössische Technische Hochschule - Swiss Federal Institute of Technology in Zürich [Zürich] (ETH Zürich), Université de Lausanne (UNIL), Laboratoire de Chimie Physique Macromoléculaire (LCPM), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Equipe NuTox (LNC - U1231) (NUTOX), Service de Médecine Néonatale [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Université de Maurice, Service de médecine foetale et foetopathologie, Service de Génétique, Hôpital de Hautepierre [Strasbourg], Department of Experimental Cardiology, Academic Medical Center - Academisch Medisch Centrum [Amsterdam] (AMC), and University of Amsterdam [Amsterdam] (UvA)-University of Amsterdam [Amsterdam] (UvA)-Heart Failure Research Center (HFRC)

Subjects
0301 basic medicine, Male, MESH: Limb Deformities, Congenital, Limb Deformities, Congenital, MESH: Phosphotransferases, [SDV.CAN]Life Sciences [q-bio]/Cancer, 030105 genetics & heredity, Biology, MESH: Founder Effect, Article, 03 medical and health sciences, Pulmonary hypoplasia, 0302 clinical medicine, Fryns syndrome, Loss of Function Mutation, Genetics, medicine, Missense mutation, Humans, Allele, [ SDV.GEN.GH ] Life Sciences [q-bio]/Genetics/Human genetics, Genetics (clinical), Loss function, Hernia, Diaphragmatic, [SDV.GEN]Life Sciences [q-bio]/Genetics, MESH: Humans, Phosphotransferases, MESH: Infant, Newborn, Infant, Newborn, Congenital diaphragmatic hernia, Facies, Infant, MESH: Loss of Function Mutation, medicine.disease, MESH: Infant, Hypoplasia, Founder Effect, MESH: Male, MESH: Hernia, Diaphragmatic, MESH: Facies, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, MESH: Gene Deletion, Female, MESH: Female, 030217 neurology & neurosurgery, Gene Deletion, Founder effect
Abstract

International audience; Fryns syndrome (FS) is a multiple malformations syndrome with major features of congenital diaphragmatic hernia, pulmonary hypoplasia, craniofacial dysmorphic features, distal digit hypoplasia, and a range of other lower frequency malformations. FS is typically lethal in the fetal or neonatal period. Inheritance is presumed autosomal recessive. Although no major genetic cause has been identified for FS, biallelic truncating variants in PIGN, encoding a component of the glycosylphosphatidylinositol (GPI)-anchor biosynthesis pathway, have been identified in a limited number of cases with a phenotype compatible with FS. Biallelic variants in PIGN, typically missense or compound missense with truncating, also cause multiple congenital anomalies-hypotonia-seizures syndrome 1 (MCAHS1). Here we report six further patients with FS with or without congenital diaphragmatic hernia and recessive loss of function PIGN alleles, including an intragenic deletion with a likely founder effect in La Réunion and other Indian Ocean islands. Our results support the hypothesis that a spectrum of phenotypic severity is associated with recessive PIGN variants, ranging from FS at the extreme end, caused by complete loss of function, to MCAHS1, in which some residual PIGN function may remain. Our data add FS resulting from PIGN variants to the catalog of inherited GPI deficiencies caused by the disruption of the GPI-anchor biosynthesis pathway.

Published
2018

14. Pediatric-onset Evans syndrome: Heterogeneous presentation and high frequency of monogenic disorders including LRBA and CTLA4 mutations

Author

Stéphane Blanche, Frédéric Rieux-Laucat, Isabelle Thuret, Olivier Alibeu, Eric Jeziorski, Pierre Quartier, Patrick Nitschke, Aude Magerus-Chatinet, Gérard Michel, Guy Leverger, Marie-Claude Stolzenberg, Geneviève de Saint-Basile, Felipe Suarez, Capucine Picard, Benedicte Neven, Nathalie Aladjidi, Nizar Mahlaoui, Eva Lévy, Alain Fischer, Isabelle Pellier, Olivier Hermine, Caroline Besnard, Judith Landman-Parker, Immunité et cancer (U932), Université Paris Descartes - Paris 5 (UPD5)-Institut Curie [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Immunogenetics of pediatric autoimmune diseases (Equipe Inserm U1163), Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Plateforme Bioinformatique [SFR Necker] (BIP-D), Structure Fédérative de Recherche Necker (SFR Necker - UMS 3633 / US24), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5), Université Sorbonne Paris Cité (USPC), CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Paris Descartes - Faculté de Médecine (UPD5 Médecine), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Centre de Référence Déficits Immunitaires Héréditaires (CEREDIH), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Service d'immuno-hématologie pédiatrique [CHU Necker], Centre Référence des Maladies Héréditaires du Métabolisme de l'Enfant et de l'Adulte [CHU Necker] (MaMEA Necker), Service de pédiatrie, d'hématologie et d'oncologie [Hôpital de La Timone - APHM], Hôpital de la Timone [CHU - APHM] (TIMONE)-Assistance Publique - Hôpitaux de Marseille (APHM), Chaire Médecine expérimentale (A. Fischer), Collège de France (CdF (institution)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Curie-Université Paris Descartes - Paris 5 (UPD5), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), Assistance Publique - Hôpitaux de Marseille (APHM)- Hôpital de la Timone [CHU - APHM] (TIMONE), Collège de France (CDF), and Collège de France (CdF)

Subjects
0301 basic medicine, Male, Evans syndrome, [SDV]Life Sciences [q-bio], Immunology, medicine.disease_cause, LRBA, Hypogammaglobulinemia, 03 medical and health sciences, Immune system, Extensive genetic screening, Autoimmune cytopenias, Immunology and Allergy, Medicine, Cytotoxic T cell, Humans, CTLA-4 Antigen, LRBA and CTLA-4 deficiencies, Preschool, Child, Purpura, Adaptor Proteins, Signal Transducing, Purpura, Thrombocytopenic, Idiopathic, business.industry, Signal Transducing, Adaptor Proteins, High-Throughput Nucleotide Sequencing, Anemia, Idiopathic, Immune dysregulation, medicine.disease, Thrombocytopenia, 3. Good health, Thrombocytopenic, 030104 developmental biology, Child, Preschool, Autoimmune lymphoproliferative syndrome, Mutation, Immune checkpoint deficiencies, Female, Anemia, Hemolytic, Autoimmune, Autoimmune hemolytic anemia, business, Hemolytic, Autoimmune
Abstract

International audience; Evans syndrome (ES) is defined by the combination of autoimmune hemolytic anemia and immune thrombocytopenia. Clinical presentation includes manifestations of immune dysregulation, found in primary immune deficiencies, autoimmune lymphoproliferative syndrome with FAS (ALPS-FAS), Cytotoxic T Lymphocyte Antigen-4 (CTLA-4) and Lipopolysaccharide-Responsive vesicle trafficking Beige-like and Anchor protein (LRBA) defects. We report the clinical history and genetic results of 18 children with ES after excluding ALPS-FAS. Thirteen had organomegaly, five lymphocytic infiltration of non-lymphoid organs, nine hypogammaglobulinemia and fifteen anomalies in lymphocyte phenotyping. Seven patients had genetic defects: three CTLA4 mutations (c.151C\textgreaterT; c.109+1092\₅68-512del; c.110-2A\textgreaterG) identified by Sanger sequencing and four revealed by Next Generation Sequencing: LRBA (c.2450+1C\textgreaterT), STAT3 gain-of-function (c.2147C\textgreaterT; c.2144C\textgreaterT) and KRAS (c.37G\textgreaterT). No feature emerged to distinguish patients with or without genetic diagnosis. Our data on pediatric-onset ES should prompt physicians to perform extensive screening for mutations in the growing pool of genes involved in primary immune deficiencies with autoimmunity.

Published
2018

15. Whole exome sequencing diagnoses the first fetal case of Bainbridge-Ropers syndrome presenting as pontocerebellar hypoplasia type 1

Author

Lucile Boutaud, Patrick Nitschke, Naima Talhi, Olivier Alibeu, Séverine Bacrot, Charlotte Mechler, Tania Attié-Bitach, Amale Ichkou, Dominique Martin-Coignard, Michel Vekemans, Sophie Thomas, Ferechté Razavi, Caroline Michot, and Philippe Roth

Subjects
0301 basic medicine, Adult, Embryology, Health, Toxicology and Mutagenesis, Genetic counseling, Pontocerebellar hypoplasia, 030105 genetics & heredity, Toxicology, Bioinformatics, Diagnosis, Differential, 03 medical and health sciences, Fetus, Exome Sequencing, Medicine, Humans, Exome sequencing, Arthrogryposis, business.industry, Syndrome, medicine.disease, Phenotype, Pediatrics, Perinatology and Child Health, Mutation (genetic algorithm), Failure to thrive, Olivopontocerebellar Atrophies, medicine.symptom, business, Developmental Biology
Abstract

BACKGROUND Bainbridge-Ropers syndrome (BRPS) is a recently identified severe disorder characterized by failure to thrive, facial dysmorphism, and severe developmental delay, caused by de novo dominant loss of function mutation in the ASXL3 gene. CASE We report here the first case of prenatal BRPS in a fetus presenting with arthrogryposis on ultrasound and for pontocerebellar hypoplasia type 1 (PCH1) following neuropathological examination. The diagnosis was done by whole exome sequencing that identified a novel de novo ASXL3 mutation. We review 29 previous published cases. DISCUSSION The fetopathological examination allowed to extend the phenotype to central nervous system and the genetic study highlights ASXL3 as a dominant gene responsible for PCH1 phenotype. Recognizing heterozygous ASXL3 mutation as a cause of prenatal PCH1 is essential for both large scale molecular analysis in the NGS era and genetic counseling.

Published
2017

16. MED13L loss-of-function variants in two patients with syndromic Pierre Robin sequence

Author

Nathalie Boddaert, Marc Bras, Maya Chopra, Stanislas Lyonnet, Patrick Nitschké, Jeanne Amiel, Christine Bole-Feysot, Christopher T. Gordon, Véronique Abadie, Myriam Oufadem, and Olivier Alibeu

Subjects
0301 basic medicine, Male, DNA Mutational Analysis, 030105 genetics & heredity, Biology, 03 medical and health sciences, Mediator, Loss of Function Mutation, Intellectual disability, Genetics, medicine, Humans, Copy-number variation, Child, Genetics (clinical), Loss function, Exome sequencing, Genetic Association Studies, Splice site mutation, Mediator Complex, Pierre Robin Syndrome, Point mutation, Brain, Infant, Sequence Analysis, DNA, medicine.disease, Magnetic Resonance Imaging, Phenotype, Child, Preschool, Female, Differential diagnosis
Abstract

We report two unrelated patients with Pierre Robin sequence (PRS) and a strikingly similar combination of associated features. Whole exome sequencing was performed for both patients. No single gene containing likely pathogenic point mutations in both patients could be identified, but the finding of an essential splice site mutation in mediator complex subunit 13 like (MED13L) in one patient prompted the investigation of copy number variants in MED13L in the other, leading to the identification of an intragenic deletion. Disruption of MED13L, encoding a component of the Mediator complex, is increasingly recognized as the cause of an intellectual disability syndrome with associated facial dysmorphism. Our findings suggest that MED13L-related disorders are a possible differential diagnosis for syndromic PRS.

Published
2017

17. Targeted Exome Sequencing Identifies

Author

Laurence, Heidet, Vincent, Morinière, Charline, Henry, Lara, De Tomasi, Madeline Louise, Reilly, Camille, Humbert, Olivier, Alibeu, Cécile, Fourrage, Christine, Bole-Feysot, Patrick, Nitschké, Frédéric, Tores, Marc, Bras, Marc, Jeanpierre, Christine, Pietrement, Dominique, Gaillard, Marie, Gonzales, Robert, Novo, Elise, Schaefer, Joëlle, Roume, Jelena, Martinovic, Valérie, Malan, Rémi, Salomon, Sophie, Saunier, Corinne, Antignac, and Cécile, Jeanpierre

Subjects
Cohort Studies, DNA-Binding Proteins, Male, Basic Research, Proto-Oncogene Proteins, Urogenital Abnormalities, DNA Mutational Analysis, Pre-B-Cell Leukemia Transcription Factor 1, Humans, Exome, Female
Abstract

Congenital anomalies of the kidney and urinary tract (CAKUT) occur in three to six of 1000 live births, represent about 20% of the prenatally detected anomalies, and constitute the main cause of CKD in children. These disorders are phenotypically and genetically heterogeneous. Monogenic causes of CAKUT in humans and mice have been identified. However, despite high-throughput sequencing studies, the cause of the disease remains unknown in most patients, and several studies support more complex inheritance and the role of environmental factors and/or epigenetics in the pathophysiology of CAKUT. Here, we report the targeted exome sequencing of 330 genes, including genes known to be involved in CAKUT and candidate genes, in a cohort of 204 unrelated patients with CAKUT; 45% of the patients were severe fetal cases. We identified pathogenic mutations in 36 of 204 (17.6%) patients. These mutations included five de novo heterozygous loss of function mutations/deletions in the PBX homeobox 1 gene (PBX1), a gene known to have a crucial role in kidney development. In contrast, the frequency of SOX17 and DSTYK variants recently reported as pathogenic in CAKUT did not indicate causality. These findings suggest that PBX1 is involved in monogenic CAKUT in humans and call into question the role of some gene variants recently reported as pathogenic in CAKUT. Targeted exome sequencing also proved to be an efficient and cost-effective strategy to identify pathogenic mutations and deletions in known CAKUT genes.

Published
2017

18. Mutations in NONO lead to syndromic intellectual disability and inhibitory synaptic defects

Author

Giovanna Bosshard, Steven J Moss, Christine Bole-Feysot, Jean-Marc Fritschy, Karine Siquier-Pernet, Patrick Nitschké, Olivier Alibeu, Ann-Kristina Fritz, Dennis Mircsof, David P. Wolfer, Florence Crestani, Markus Rudin, Steven A. Brown, Nathalie Boddaert, Rochelle M. Hines, Arnold Munnich, Jeanne Amiel, Petra Seebeck, Matej Žnidarič, Christina Koester, Nicolas Cagnard, Ludmila Gaspar, Maéva Langouët, Shiva K. Tyagarajan, Laurence Colleaux, Aileen Schröter, Marlène Rio, Sébastien Moutton, Katrina Prescott, Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris Descartes - Paris 5 (UPD5), Université Sorbonne Paris Cité (USPC), CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Service de Génétique Médicale [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Plateforme de génomique [SFR Necker], Structure Fédérative de Recherche Necker (SFR Necker - UMS 3633 / US24), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Plateforme Bioinformatique [SFR Necker] (BIP-D), Institute of Anatomy, Universität Zürich [Zürich] = University of Zurich (UZH), Service de radiologie pédiatrique [CHU Necker], Department of Clnical Genetics, Chapel Allerton Hospital, Institute of Pharmacology and Toxicology [Zurich], University of Zurich, Brown, Steven A, Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Necker - Enfants Malades [AP-HP], Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Assistance publique - Hôpitaux de Paris (AP-HP) (APHP), University of Zürich [Zürich] (UZH), and Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)

Subjects
Male, Protein family, Adolescent, 10050 Institute of Pharmacology and Toxicology, RNA-binding protein, 610 Medicine & health, Mice, Transgenic, Biology, Inhibitory postsynaptic potential, Article, Mice, Nuclear Matrix-Associated Proteins, Cellular neuroscience, Intellectual Disability, Intellectual disability, medicine, Transcriptional regulation, Animals, Humans, 10237 Institute of Biomedical Engineering, Cells, Cultured, ComputingMilieux_MISCELLANEOUS, Genetics, Gephyrin, General Neuroscience, 2800 General Neuroscience, Brain, RNA-Binding Proteins, Neural Inhibition, medicine.disease, Pedigree, DNA-Binding Proteins, Mice, Inbred C57BL, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, Mutation, Synapses, biology.protein, GABAergic, 570 Life sciences, biology, Octamer Transcription Factors, Neuroscience
Abstract

The NONO protein has been characterized as an important transcriptional regulator in diverse cellular contexts. Here we show that loss of NONO function is a likely cause of human intellectual disability and that NONO-deficient mice have cognitive and affective deficits. Correspondingly, we find specific defects at inhibitory synapses, where NONO regulates synaptic transcription and gephyrin scaffold structure. Our data identify NONO as a possible neurodevelopmental disease gene and highlight the key role of the DBHS protein family in functional organization of GABAergic synapses.

Published
2015

19. Cakutome, a high-throughput tool for molecular diagnosis and identification of novel causative genes for CAKUT patients

Author

Cécile Jeanpierre, Olivier Alibeu, Charline Henry, E. Schaeffer, Christine Bole-Feysot, Robert Novo, Rémi Salomon, Christine Pietrement, R. Campait, Jelena Martinovic, Joelle Roume, Patrick Nitschke, Marie Gonzales, L. De Tomasi, Dominique Gaillard, Laurence Heidet, Sophie Saunier, Corinne Antignac, Cécile Fourrage, and Vincent Morinière

Subjects
Sanger sequencing, Biallelic Mutation, Genetics, Mutation, Candidate gene, Genetic heterogeneity, Biology, HNF1B, medicine.disease_cause, symbols.namesake, Pediatrics, Perinatology and Child Health, symbols, medicine, Copy-number variation, Exome sequencing
Abstract

Introduction Congenital anomalies of the kidney and urinary tract (CAKUT) are a major cause of renal failure in children. CAKUT are phenotypically and genetically heterogeneous and more than 50 genes have been reported as mutated in patients. The most frequently mutated genes are those encoding transcription factors HNF1B, PAX2, EYA1 and SIX1. Most of the other genes are only mutated in a few patients and their implication is sometimes elusive. Methods We developped a targeted exome sequencing strategy («CAKUTOME», sureselect Agilent) focusing on 330 genes, including known validated or likely causative CAKUT genes, as well as candidate genes. A total of 215 unrelated patients were analysed, including 50 who had previously been tested for HNF1B, PAX2, EYA1 and/or SIX1 mutations by Sanger sequencing. Results/discussion/conclusion This approach proved to be an efficient and cost-effective strategy to identify pathogenic mutations and copy number variations in known CAKUT genes. The 25/165 rate of mutation we identified in HNF1B, PAX2, EYA1 or SIX1 is similar to the one obtained by Sanger sequencing. In addition, we identified heterozygous mutations in ANOS1 (Kallmann syndrome), GATA3 (hypoparathyroidism, deafness and kidney disease), or CHD7 (Charge syndrome), and biallelic mutation in KIF14 in 2, 3, 1 and 2 cases, respectively. Our data also led to the identification of a novel CAKUT gene, the mutation/deletion of which affected 5 unrelated cases in the cohort. Moreover, we identified variants in several other genes never reported as mutated in CAKUT patients, whose pathogenicity is being tested. Finally, no relevant variant was identified in 40 % of our series. Although mutations in gene(s), non-coding regions or microRNAs not targeted in our CAKUTOME could be involved in some of these cases, complex inheritance, somatic events, and/or environmental factors or epigenetic mechanisms likely explain this large fraction of cases.

Published
2017

20. CE-SSCP and CE-FLA, simple and high-throughput alternatives for fungal diversity studies

Author

Jérôme Gury, Lucie Zinger, Lucile Sage, François Pompanon, Roberto A. Geremia, Olivier Alibeu, Delphine Rioux, Ludovic Gielly, Laboratoire d'Ecologie Alpine (LECA), and Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])

Subjects
microbial, t-rflp, Biodiversity, registered designation, chemistry.chemical_compound, Molecular marker, DNA, Fungal, high-throughput, Polymorphism, Single-Stranded Conformational, Soil Microbiology, [SDV.EE]Life Sciences [q-bio]/Ecology, environment, 0303 health sciences, Ecology, Community structure, keratitis, gradient gel-electrophoresis, community structure, origin salers, Polymorphism, Restriction Fragment Length, Microbiology (medical), ribosomal-rna genes, [SDV.BID]Life Sciences [q-bio]/Biodiversity, Biology, Sensitivity and Specificity, Microbiology, soil, cheese, 03 medical and health sciences, Capillary electrophoresis, CE-FLA, Ecosystem, CE-SSCP, Molecular Biology, 030304 developmental biology, 030306 microbiology, Fungi, Electrophoresis, Capillary, Reproducibility of Results, Single-strand conformation polymorphism, 15. Life on land, DNA Fingerprinting, soil bacterial, genomic DNA, capillary-electrophoresis, Agronomy, chemistry, mycotic, Soil water, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, Strand-conformation polymorphism
Abstract

International audience; Fungal communities are key components of soil, but the study of their ecological significance is limited by a lack of appropriated methods. For instance, the assessment of fungi occurrence and spatio-temporal variation in soil requires the analysis of a large number of samples. The molecular signature methods provide a useful tool to monitor these microbial communities and can be easily adapted to capillary electrophoresis (CE) allowing high-throughput studies. Here we assess the suitability of CE-FLA (Fragment Length Polymorphism, denaturing conditions) and CE-SSCP (Single-Stranded Conformation Polymorphism, native conditions) applied to environmental studies since they require a short molecular marker and no post-PCR treatments. We amplified the ITSI region from 22 fungal strains isolated from an alpine ecosystem and from total genomic DNA of alpine and infiltration basin soils. The CE-FLA and CE-SSCP separated 17 and 15 peaks respectively from a mixture of 19 strains. For the alpine soil-metagenomic DNA, the FLA displayed more peaks than the SSCP and the converse result was found for infiltration basin sediments. We concluded that CE-FLA and CE-SSCP of ITS I region provided complementary information. In order to improve CE-SSCP sensitivity. we tested its resolution according to migration temperature and found 32 degrees C to be optimal. Because of their simplicity, quickness and reproducibility. we found that these two methods were promising for high-throughput studies of soil fungal communities. (c) 2007 Elsevier B.V All rights reserved.

Published
2008

21. Mutations in SNRPB, encoding components of the core splicing machinery, cause cerebro-costo-mandibular syndrome

Author

Didier Lacombe, Séverine Bacrot, Niklas Feldhahn, Florence Petit, Mathilde Doyard, Daphné Lehalle, Patrick Nitschke, Arnold Munnich, Marie-Paule Vazquez, Olivier Alibeu, Céline Huber, Sandrine Marlin, and Valérie Cormier-Daire

Subjects
Adult, Male, Spliceosome, Heterozygote, Adolescent, Micrognathism, Mutation, Missense, Ribs, Biology, snRNP Core Proteins, Exon, symbols.namesake, Young Adult, Intellectual Disability, Genetics, Humans, Gene, Genetics (clinical), Exome sequencing, Genetic Association Studies, Sanger sequencing, Base Sequence, Alternative splicing, Molecular biology, Child, Preschool, RNA splicing, symbols, Small nuclear ribonucleoprotein
Abstract

Cerebro-costo-mandibular syndrome (CCMS) is a developmental disorder characterized by the association of Pierre Robin sequence and posterior rib defects. Exome sequencing and Sanger sequencing in five unrelated CCMS patients revealed five heterozygous variants in the small nuclear ribonucleoprotein polypeptides B and B1 (SNRPB) gene. This gene includes three transcripts, namely transcripts 1 and 2, encoding components of the core spliceosomal machinery (SmB' and SmB) and transcript 3 undergoing nonsense-mediated mRNA decay. All variants were located in the premature termination codon (PTC)-introducing alternative exon of transcript 3. Quantitative RT-PCR analysis revealed a significant increase in transcript 3 levels in leukocytes of CCMS individuals compared to controls. We conclude that CCMS is due to heterozygous mutations in SNRPB, enhancing inclusion of a SNRPB PTC-introducing alternative exon, and show that this developmental disease is caused by defects in the splicing machinery. Our finding confirms the report of SNRPB mutations in CCMS patients by Lynch et al. (2014) and further extends the clinical and molecular observations.

Published
2014

22. Mutations of CEP83 Cause Infantile Nephronophthisis and Intellectual Disability

Author

Pauline Krug, Emilie Filhol, Robert Novo, Amandine Frigo, Olivier Alibeu, Daniela A. Braun, Ce´cile Masson, Karine Brochard, Lilya Belkacem, Alexandre Benmerah, Christine Pietrement, Marion Failler, Markus Schueler, Re´mi Salomon, Heon Yung Gee, Jonathan D. Porath, Bruno Hurault de Ligny, Joon Kim, Marie-Claire Gubler, Jan Halbritter, Edgar A. Otto, Kwangsic Joo, Friedhelm Hildebrandt, Hülya Kayserili, Corinne Antignac, and Sophie Saunier

Subjects
Central Nervous System, Male, Centriole, Biology, Ciliopathies, 03 medical and health sciences, Exon, 0302 clinical medicine, Nephronophthisis, Ciliogenesis, Report, Intellectual Disability, Genetics, medicine, Humans, Genetics(clinical), Cilia, Genetics (clinical), Exome sequencing, Alleles, 030304 developmental biology, Centrioles, 0303 health sciences, Cilium, Infant, Exons, Kidney Diseases, Cystic, Orofaciodigital Syndromes, medicine.disease, Ciliopathy, Child, Preschool, Mutation, Female, Microtubule-Associated Proteins, 030217 neurology & neurosurgery
Abstract

Ciliopathies are a group of hereditary disorders associated with defects in cilia structure and function. The distal appendages (DAPs) of centrioles are involved in the docking and anchoring of the mother centriole to the cellular membrane during ciliogenesis. The molecular composition of DAPs was recently elucidated and mutations in two genes encoding DAPs components (CEP164/NPHP15, SCLT1) have been associated with human ciliopathies, namely nephronophthisis and orofaciodigital syndrome. To identify additional DAP components defective in ciliopathies, we independently performed targeted exon sequencing of 1,221 genes associated with cilia and 5 known DAP protein-encoding genes in 1,255 individuals with a nephronophthisis-related ciliopathy. We thereby detected biallelic mutations in a key component of DAP-encoding gene, CEP83, in seven families. All affected individuals had early-onset nephronophthisis and four out of eight displayed learning disability and/or hydrocephalus. Fibroblasts and tubular renal cells from affected individuals showed an altered DAP composition and ciliary defects. In summary, we have identified mutations in CEP83, another DAP-component-encoding gene, as a cause of infantile nephronophthisis associated with central nervous system abnormalities in half of the individuals.

Published
2014

23. Corrigendum to 'CE-SSCP and CE-FLA, simple and high-throughput alternatives for fungal diversity studies' [Journal of Microbiological Methods 72 (2008) 42–53]

Author

François Pompanon, Roberto A. Geremia, Ludovic Gielly, Olivier Alibeu, Delphine Rioux, Jérôme Gury, Lucile Sage, Lucie Zinger, Laboratoire d'Ecologie Alpine (LECA), and Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])

Subjects
2. Zero hunger, 0106 biological sciences, Microbiology (medical), [SDV.EE]Life Sciences [q-bio]/Ecology, environment, business.industry, Library science, 04 agricultural and veterinary sciences, English language, [SDV.BID]Life Sciences [q-bio]/Biodiversity, 15. Life on land, Biology, 01 natural sciences, Microbiology, Biotechnology, Fungal Diversity, 040103 agronomy & agriculture, 0401 agriculture, forestry, and fisheries, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, business, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 010606 plant biology & botany
Abstract

The authors are indebted to D. A. Monier and C. Miquel (LECA) for expert technical help, to P. Bedell and C. Delolme (LSE, Lyon) and P. Choler for soil samples and A. Cleary for correcting the English language. F. Pompanon was supported by the French Institut National de la Recherche Agronomique. This work was supported by the ANR Project MICROALPES (ANR-06-BLAN-0301), program ECCO EMMAUS, and CNRS funding.

Published
2008

24. Phylogenetic analysis of the Aspergillus niger aggregate in relation to feruloyl esterase activity

Author

Joëlle Dupont, Laurence Lesage-Meessen, Frédéric Giraud, David Navarro, Marcel Asther, Françoise Seigle-Murandi, Mireille Haon, Ourdia Bouzid, Olivier Alibeu, Lucile Sage, Taxonomie - Collections (TC), Centre National de la Recherche Scientifique (CNRS), Biodiversité et Biotechnologie Fongiques (BBF), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-École Centrale de Marseille (ECM), Laboratoire d'Ecologie Alpine (LECA), Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Université Joseph Fourier - Grenoble 1 (UJF)-Université Grenoble Alpes (UGA), Muséum national d'Histoire naturelle (MNHN)-Centre National de la Recherche Scientifique (CNRS), Université Joseph Fourier - Grenoble 1 (UJF)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Centre National de la Recherche Scientifique (CNRS), Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), and École Centrale de Marseille (ECM)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA)

Subjects
Molecular Sequence Data, beta-tubulin, ochratoxin-a, [SDV.BID]Life Sciences [q-bio]/Biodiversity, phylogeny, Microbiology, aflp, Fungal Proteins, 03 medical and health sciences, Phylogenetics, Feruloyl esterase, Tubulin, Botany, DNA, Ribosomal Spacer, acid esterases, feruloyl esterase, sugar-beet pulp, DNA, Fungal, Molecular Biology, 030304 developmental biology, black aspergilli, degradation, [SDV.EE]Life Sciences [q-bio]/Ecology, environment, 0303 health sciences, Fungal protein, Phylogenetic tree, biology, tubingensis, 030306 microbiology, Aspergillus niger, General Medicine, Fungi imperfecti, Sequence Analysis, DNA, Length polymorphism analysis, biology.organism_classification, classification, Sugar beet, Amplified fragment length polymorphism, ITS, section nigri, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, Carboxylic Ester Hydrolases, Polymorphism, Restriction Fragment Length
Abstract

International audience; Species of the Aspergillus niger aggregate are known to produce feruloyl esterases, enzymes involved in the degradation of cell wall polymers. However, species delineation is difficult in these fungi. We combined AFLP analysis with ITS rDNA and beta-tubulin sequencing to characterize the isolates of this aggregate in terms of feruloyl esterase production. A preliminary re-examination of isolates based on comparison of ITS rDNA and beta-tubulin sequences with those of typical taxa deposited in international collections led us to re-identify the isolates as members of the species A. niger, A. foetidus and A. tubingensis. Molecular clustering based on beta-tubulin data and AFLP analysis showed that the strains of A. niger formed a homogenous phylogenetic group distinguished by either zero or type A feruloyl esterase activity, while strains A. foetidus and A. tubingensis exhibited type B feruloyl esterase activity when grown on sugar beet pulp. (c) 2007 Elsevier Masson SAS. All rights reserved.

Published
2007

25. PAXX and Xlf interplay revealed by impaired CNS development and immunodeficiency of double KO mice

Author

Ana Cumano, Benoit Roch, Laetitia Kermasson, Ramy Elsaid, Vincent Abramowski, Jean-Paul Moussu, Olivier Etienne, Aurélie Berland, Jean-Pierre de Villartay, Stefania Musilli, Patrick Revy, Junjie Yang, François D. Boussin, Karelia Lipson-Ruffert, Imagine - Institut des maladies génétiques (IMAGINE - U1163), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire de Radiopathologie (LRP), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Lymphopoïèse (Lymphopoïèse (UMR_1223 / U1223 / U-Pasteur_4)), Institut Pasteur [Paris] (IP)-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Cellule Pasteur, Université Paris Diderot - Paris 7 (UPD7)-PRES Sorbonne Paris Cité, Transgenèse et archivage d'animaux modèles (TAAM), Centre National de la Recherche Scientifique (CNRS), This work was supported by institutional grants from INSERM, the Institut National du Cancer (PLBIO16-280), the Agence Nationale de la Recherche ('Investissement d’Avenir' program ANR-10-IAHU-01), and by grants from La Ligue Nationale contre le Cancer (Equipe Labellisée LA LIGUE), Pasteur Institute, ANR (Program REVIVE and grant Twothyme) to AC., We thank Catherine Cailleau (SEAT) for exceptional care of our mouse lines, Sophie Berissi for help in immunohistochemistry and Olivier Alibeu, Aurore Pouliet, and Christine Bole-Feyssot (Genomic facility, Imagine Institute) for their assistance in NGS, ANR-10-LABX-0073,REVIVE,Stem Cells in Regenerative Biology and Medicine(2010), ANR-14-CE11-0022,Twothyme,Deux progéniteurs hématopoïétiques différents établissent le compartiment de lymphocytes T: tester un nouveau paradigme du développement T.(2014), ANR-10-IAHU-0001,Imagine,Institut Hospitalo-Universitaire Imagine(2010), Vougny, Marie-Christine, Laboratoires d'excellence - Stem Cells in Regenerative Biology and Medicine - - REVIVE2010 - ANR-10-LABX-0073 - LABX - VALID, Appel à projets générique - Deux progéniteurs hématopoïétiques différents établissent le compartiment de lymphocytes T: tester un nouveau paradigme du développement T. - - Twothyme2014 - ANR-14-CE11-0022 - Appel à projets générique - VALID, Instituts Hospitalo-Universitaires - Institut Hospitalo-Universitaire Imagine - - Imagine2010 - ANR-10-IAHU-0001 - IAHU - VALID, Université Paris Diderot - Paris 7 (UPD7)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Institut Pasteur [Paris]-Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM)

Subjects
0301 basic medicine, Central Nervous System, DNA End-Joining Repair, [SDV.IMM] Life Sciences [q-bio]/Immunology, Biology, medicine.disease_cause, 03 medical and health sciences, Mice, Immune system, medicine, Animals, Lymphocytes, Molecular Biology, Immunodeficiency, Loss function, chemistry.chemical_classification, Mice, Knockout, DNA ligase, Mutation, Original Paper, Genetic interaction, Immunologic Deficiency Syndromes, Cell Biology, DNA repair protein XRCC4, medicine.disease, Acquired immune system, Phenotype, Cell biology, Resin Cements, DNA-Binding Proteins, Mice, Inbred C57BL, 030104 developmental biology, chemistry, Immunology, [SDV.IMM]Life Sciences [q-bio]/Immunology
Abstract

International audience; The repair of DNA double-stranded breaks (DNAdsb) through non-homologous end joining (NHEJ) is a prerequisite for the proper development of the central nervous system and the adaptive immune system. Yet, mice with Xlf or PAXX loss of function are viable and present with very mild immune phenotypes, although their lymphoid cells are sensitive to ionizing radiation attesting for the role of these factors in NHEJ. In contrast, we show here that mice defective for both Xlf and PAXX are embryonically lethal owing to a massive apoptosis of post-mitotic neurons, a situation reminiscent to XRCC4 or DNA Ligase IV KO conditions. The development of the adaptive immune system in Xlf(-/-)PAXX(-/-) E18.5 embryos is severely affected with the block of B- and T-cell maturation at the stage of IgH and TCRβ gene rearrangements, respectively. This damaging phenotype highlights the functional nexus between Xlf and PAXX, which is critical for the completion of NHEJ-dependent mechanisms during mouse development.Cell Death and Differentiation advance online publication, 27 October 2017; doi:10.1038/cdd.2017.184.

Published
2017

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