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Item does not contain fulltext BACKGROUND: Cohesinopathies are rare neurodevelopmental disorders arising from a dysfunction in the cohesin pathway, which enables chromosome segregation and regulates gene transcription. So far, eight genes from this pathway have been reported in human disease. STAG1 belongs to the STAG subunit of the core cohesin complex, along with five other subunits. This work aimed to identify the phenotype ascribed to STAG1 mutations. METHODS: Among patients referred for intellectual disability (ID) in genetics departments worldwide, array-comparative genomic hybridisation (CGH), gene panel, whole-exome sequencing or whole-genome sequencing were performed following the local diagnostic standards. RESULTS: A mutation in STAG1 was identified in 17 individuals from 16 families, 9 males and 8 females aged 2-33 years. Four individuals harboured a small microdeletion encompassing STAG1; three individuals from two families had an intragenic STAG1 deletion. Six deletions were identified by array-CGH, one by whole-exome sequencing. Whole-exome sequencing found de novo heterozygous missense or frameshift STAG1 variants in eight patients, a panel of genes involved in ID identified a missense and a frameshift variant in two individuals. The 17 patients shared common facial features, with wide mouth and deep-set eyes. Four individuals had mild microcephaly, seven had epilepsy. CONCLUSIONS: We report an international series of 17 individuals from 16 families presenting with syndromic unspecific ID that could be attributed to a STAG1 deletion or point mutation. This first series reporting the phenotype ascribed to mutation in STAG1 highlights the importance of data sharing in the field of rare disorders.

International audience; Neutropenia can be qualified as congenital when of neonatal onset or when associated with extra-hematopoietic manifestations. Overall, 30% of patients with congenital neutropenia (CN) remain without a molecular diagnosis after a multidisciplinary consultation and tedious diagnostic strategy. In the rare situations when neutropenia is identified and associated with intellectual disability (ID), there are few diagnostic hypotheses to test. This retrospective multicenter study reports on a clinically heterogeneous cohort of 10 unrelated patients with CN associated with ID and no molecular diagnosis prior to whole-exome sequencing (WES). WES provided a diagnostic yield of 40% (4/10). The results suggested that in many cases neutropenia and syndromic manifestations could not be assigned to the same molecular alteration. Three sub-groups of patients were highlighted: (i) severe, symptomatic chronic neutropenia, detected early in life, and related to a known mutation in the CN spectrum (ELANE); (ii) mild to moderate benign intermittent neutropenia, detected later, and associated with mutations in genes implicated in neurodevelopmental disorders (CHD2, HUWE1); and (iii) moderate to severe intermittent neutropenia as a probably undiagnosed feature of a newly reported syndrome (KAT6A). Unlike KAT6A, which seems to be associated with a syndromic form of CN, the other reported mutations may not explain the entire clinical picture. Although targeted gene sequencing can be discussed for the primary diagnosis of severe CN, we suggest that performing WES for the diagnosis of disorders associating CN with ID will not only provide the etiological diagnosis but will also pave the way towards personalized care and follow-up. (C) 2016 Wiley Periodicals, Inc., (C) 2016 John Wiley & Sons, Ltd

International audience; Self-renewal and differentiation of pluripotent murine embryonic stem cells (ESCs) is regulated by extrinsic signaling pathways. It is less clear whether cellular metabolism instructs developmental progression. In an unbiased genome-wide CRISPR/Cas9 screen, we identified components of a conserved amino-acid-sensing pathway as critical drivers of ESC differentiation. Functional analysis revealed that lysosome activity, the Ragulator protein complex, and the tumor-suppressor protein Folliculin enable the Rag GTPases C and D to bind and seclude the bHLH transcription factor Tfe3 in the cytoplasm. In contrast, ectopic nuclear Tfe3 represses specific developmental and metabolic transcriptional programs that are associated with peri-implantation development. We show differentiation-specific and non-canonical regulation of Rag GTPase in ESCs and, importantly, identify point mutations in a Tfe3 domain required for cytoplasmic inactivation as potentially causal for a human developmental disorder. Our work reveals an instructive and biomedically relevant role of metabolic signaling in licensing embryonic cell fate transitions.

Cohen Syndrome (CS) is a rare autosomal recessive disease caused by variations in the gene coding for the vacuolar tri-vacuolar protein 13B (VPS13B or COH1). CS patients have common features including typical facial appearance, neutropenia, abnormal trunk fat distribution, microcephaly, myopia and retinal damage. My thesis project aimed to characterize the ophthalmologic phenotype of a mouse model Vps13bEx3/Ex3 to determine the molecular mechanisms involved in the development of CS retinopathy.First, we showed that Vps13bEx3/Ex3 mice develop a cataract between 2 and 3 months. Disorganization of the fibrous cells of the crystalline lens and their differentiation into mesenchymal cells were observed. At a more advanced stage, the external membrane of the crystalline lens is ruptured and the cortical and nuclear part of the crystalline lens separates. After the onset of cataract, the retina shows deformations characterized by thinning and deformation of its various layers which can lead to retinal folds 50 to 500µm long. There is also a strong proliferation of microglial cells as well as astrogliosis. The appearance of cataract in our model has led to an exhaustive review of the literature showing that cataract is present in 85% of patients at 40 years old and not related to retinopathy.We then showed a modification of the ocular phenotype in the presence of the Crb1Rd8 mutation. Crb1 is a transmembrane protein whose Rd8 variation leads to retinal disorganization. We have created cohorts of all possible Vps13b/Crb1 genotypes. Our preliminary analyses show early retinal dystrophy in mice Vps13bEx3/Ex3 Crb1Rd8/+, suggesting that Vps13b could be a Crb1 modifier gene. Molecular mechanisms that could lead to the appearance of SC retinopathy have been studied in Vps13bEx3/Ex3Crb1+/+ mice. The structure of the retina and these different cell populations were studied but did not reveal any abnormalities before the onset of the cataract. However, the study of inflammatory mechanisms showed an increase in sensitivity to IL6 in the retina of our model. This mechanism could be involved in the development of SC retinopathy. As the ocular phenotype is strongly decreased in mice Vps13bEx3/Ex3 Crb1+/+,the impact of environmental factors, especially light, has also been studied and shows that when subjected to high light intensity, mice develop retinopathies.In parallel, we have studied the ophthalmological characteristics of our Dijon cohort mainly using OCT. This study reveals (i) that macular edema is a frequent symptom of SC which remains relatively stable over time and (ii) that the onset of retinopathy is independent of that of cataract.Consequently, my PhD work has allowed to characterize the ophthalmic attacks of our Vps13bEx3/Ex3 mouse model and will allow to deepen the mechanistic knowledge of retinopathy and cataract of the CS, and could allow to develop a therapeutic target or prevention strategies for these ophthalmic lesions., Le Syndrome de Cohen (SC)(OMIM 216550) est une maladie autosomique récessive rare due à des variations dans le gène codant pour la protéine de tri vacuolaire 13B (VPS13B ou COH1). Les patients possèdent des caractéristiques cliniques typiques : une dysmorphie faciale, une neutropénie, une répartition anormale des graisses au niveau du tronc, une microcéphalie associée à une déficience intellectuelle, une myopie et des atteintes rétiniennes. Mon projet de thèse a eu pour but de caractériser le phénotype ophtalmologique du modèle murin du SC (Vps13bEx3/Ex3) afin de déterminer les mécanismes moléculaires impliqués dans la mise en place de la rétinopathie du SC.Nous avons montré que les souris Vps13bEx3/Ex3 développent une cataracte entre 2 et 3 mois d’âge, avec une désorganisation des cellules fibreuses du cristallin et leur différenciation en cellules mésenchymateuses. À un stade plus avancé, la membrane externe du cristallin est rompue et les parties corticale et nucléaire du cristallin se séparent. Après l’apparition de la cataracte, la rétine présente des déformations se caractérisant par un amincissement et une déformation de ses différentes couches pouvant engendrer des plis rétiniens de 50 à 500µm de long. Il y a également une forte prolifération des cellules microgliales ainsi qu’une astrogliose. La découverte de la cataracte dans notre modèle a conduit à une revue exhaustive de la littérature permettant de mettre en évidence que la cataracte est présente chez 85% des patients à 40 ans et sans lien avec la rétinopathie.Nous avons ensuite montré une modification du phénotype oculaire en présence de la mutation Crb1Rd8. Crb1 est une protéine transmembranaire dont la variation Rd8 entraine une désorganisation de la rétine. Nous avons créé des cohortes de tous les génotypes Vps13b/Cr1 possibles. Nos analyses préliminaires montrent une dystrophie rétinienne précoce chez les souris Vps13bEx3/Ex3 Crb1Rd8/+, suggérant que Vps13b pourrait être un gène modificateur de Crb1. Les mécanismes moléculaires pouvant conduire à l’apparition de la rétinopathie du SC ont été étudiés sur des souris Vps13bEx3/Ex3Crb1+/+. La structure de la rétine et ses différentes populations cellulaires ont été étudiées mais n’ont pas révélé d’anomalies avant l’apparition de la cataracte. Cependant l’étude des mécanismes inflammatoires a montré une augmentation de la sensibilité à IL6 dans la rétine de notre modèle. Ce mécanisme pourrait être impliqué dans la mise en place de la rétinopathie. Le phénotype oculaire étant fortement diminué chez les souris Vps13bEx3/Ex3 Crb1+/+, l’impact des facteurs environnementaux, et en particulier la lumière, a également été étudié et montre que soumises à de fortes intensités lumineuses, les souris développent des rétinopathies.En parallèle, nous avons étudié les caractéristiques ophtalmologiques des patients de notre cohorte principalement à l’aide d’OCT. Cette étude révèle (i) que l’œdème maculaire est un symptôme fréquent du SC qui reste relativement stable dans le temps et (ii) que l’apparition de la rétinopathie est indépendante de celle de la cataracte.Ainsi mes travaux de doctorat ont permis de caractériser les atteintes ophtalmiques de notre modèle murin Vps13bEx3/Ex3. L’étude des mécanismes cellulaires impliqués dans la rétinopathie et la cataracte du SC doit être approfondie afin de permettre de trouver une cible thérapeutique ou des stratégies de prévention pour ces atteintes ophtalmiques.

Oral-facial-digital syndromes (OFDS) are characterized by the association of oral, facial and digital anomalies. The different modes of inheritance and additional features lead to clinically delineate 13 subtypes. For a long time, only the OFD1 gene, responsible for OFDI subtype and coding for a centrosomal protein, has been known, suggesting the involvement of the primary cilium in OFDS. Mutations have recently been reported in the TMEM216, DDX59, SCLT1, TBC1D32 and TCTN3 genes in anecdotic cases. To identify new genes involved in OFDS, we performed whole-exome sequencing in 24 patients. In 14/24 cases, we identified 5 novel genes (C2CD3, TMEM107, INTU, KIAA0753, IFT57), enlarged the clinical spectrum of OFDS of 3 known genes responsible for other ciliopathies (C5orf42, TMEM138, TMEM231) and confirmed the involvement of 3 known genes in OFDS (OFD1, DDX59, WDPCP). Functional studies demonstrated the involvement of the centriolar growth, the transition zone and the intraflagellar transport, through the characterization of 3 major protein complexes: the KIAA0753/OFD1/FOPNL complex controlling the centriole elongation, the MKS module (TMEM107/TMEM231/TMEM216), an essential component of the transition zone, and the CPLANE complex (INTU/FUZ/WDPCP) enabling in the IFT-A assembly. We demonstrated the large clinical and genetic heterogeneity of OFDS, yielding the initial classification in 13 subtypes obsolete, extending the number of 15 causal genes, and confirming OFDS as a new full subgroup of ciliopathies., Les syndromes oro-facio-digitaux (OFD) sont caractérisés par la présence d'une atteinte orale, faciale et digitale et classés en 13 sous-types. Pendant longtemps seul le gène OFD1, responsable du type I et codant pour une protéine centrosomale et du corps basal, était principalement connu, faisant suspecter l’implication du cil primaire dans les syndromes OFD. Des mutations ont été rapportées plus récemment dans les gènes TMEM216, DDX59, SCLT1, TBC1D32 et TCTN3 chez un ou deux patients. Dans le but d’identifier de nouveaux gènes impliqués dans ces syndromes, nous avons réalisé une stratégie de séquençage haut débit d’exome chez 24 patients. Cette stratégie a permis d’identifier 5 nouveaux gènes (C2CD3, TMEM107, INTU, KIAA0753, IFT57), d'étendre aux syndromes OFD le spectre clinique de 3 gènes déjà connus dans d'autres ciliopathies (C5orf42, TMEM138, TMEM231) et de confirmer l'implication de 3 gènes déjà connus comme responsable de syndrome OFD (OFD1, DDX59, WDPCP). Les analyses fonctionnelles ont montré l’implication de la croissance centriolaire, de la zone de transition et du transport intraflagellaire, avec notamment la caractérisation de 3 complexes protéiques principaux : le complexe ternaire KIAA0753/OFD1/FOPNL, régulant la croissance centriolaire, le complexe MKS (TMEM107, TMEM231, TMEM216) constituant majeur de la zone de transition et le complexe CPLANE (INTU/FUZ/WDPCP), favorisant l’assemblage des protéines périphériques du complexe de transport intraflagellaire IFT-A. En conclusion, cette étude, la plus importante consacrée aux syndromes OFD, démontre la très grande hétérogénéité clinique et génétique de ces syndromes, de même que de nombreux allélismes avec d’autres ciliopathies. Elle étend à 15 le nombre de gènes causaux, rendant la classification clinique initiale totalement obsolète et permettant de considérer les syndromes OFD comme un nouveau sous-groupe de ciliopathies à part entière.