Publications by authors named "Sylvain Hanein"

51 Publications

High Diagnostic Yield of Targeted Next-Generation Sequencing in a Cohort of Patients With Congenital Hypothyroidism Due to Dyshormonogenesis.

Front Endocrinol (Lausanne) 2020 22;11:545339. Epub 2021 Feb 22.

INSERM U1016, Cochin Institute, Faculté de Médecine, Université Paris Descartes, Sorbonne Paris Cité, Paris, France.

Objective: To elucidate the molecular cause in a well-characterized cohort of patients with Congenital Hypothyroidism (CH) and Dyshormonogenesis (DH) by using targeted next-generation sequencing (TNGS).

Study Design: We studied 19 well-characterized patients diagnosed with CH and DH by targeted NGS including genes involved in thyroid hormone production. The pathogenicity of novel mutations was assessed based on prediction tool results, functional studies when possible, variant location in important protein domains, and a review of the recent literature.

Results: TNGS with variant prioritization and detailed assessment identified likely disease-causing mutations in 10 patients (53%). Monogenic defects most often involved , followed by , , and and were usually homozygous or compound heterozygous. Our review shows the importance of the detailed phenotypic description of patients and accurate analysis of variants to provide a molecular diagnosis.

Conclusions: In a clinically well-characterized cohort, TNGS had a diagnostic yield of 53%, in accordance with previous studies using a similar strategy. mutations were the most common genetic defect. TNGS identified gene mutations causing DH, thereby providing a rapid and cost-effective genetic diagnosis in patients with CH due to DH.
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http://dx.doi.org/10.3389/fendo.2020.545339DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7937947PMC
May 2021

Whole Locus Sequencing Identifies a Prevalent Founder Deep Intronic Pathologic Variant in the French Leber Congenital Amaurosis Cohort.

Genes (Basel) 2021 02 18;12(2). Epub 2021 Feb 18.

Laboratory of Genetics in Ophthalmology (LGO), INSERM UMR1163, Institute of Genetics Diseases, Imagine and Paris Descartes University, 75015 Paris, France.

Leber congenital amaurosis (LCA) encompasses the earliest and most severe retinal dystrophies and can occur as a non-syndromic or a syndromic disease. Molecular diagnosis in LCA is of particular importance in clinical decision-making and patient care since it can provide ocular and extraocular prognostics and identify patients eligible to develop gene-specific therapies. Routine high-throughput molecular testing in LCA yields 70%-80% of genetic diagnosis. In this study, we aimed to investigate the non-coding regions of one non-syndromic LCA gene, , in a series of six families displaying one single disease allele after a gene-panel screening of 722 LCA families which identified 26 biallelic families. Using trio-based high-throughput whole locus sequencing (WLS) for second disease alleles, we identified a founder deep intronic mutation (NM_020366.3:c.1468-128T>G) in 3/6 families. We employed Sanger sequencing to search for the pathologic variant in unresolved LCA cases (106/722) and identified three additional families (two homozygous and one compound heterozygous with the NM_020366.3:c.930+77A>G deep intronic change). This makes the c.1468-128T>G the most frequent disease allele (8/60, 13%) in our cohort. Studying patient lymphoblasts, we show that the pathologic variant creates a donor splice-site and leads to the insertion of the pseudo-exon in the mRNA, which we were able to hamper using splice-switching antisense oligonucleotides (AONs), paving the way to therapies.
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http://dx.doi.org/10.3390/genes12020287DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7922592PMC
February 2021

Low Prevalence of GSC Gene Mutations in a Large Cohort of Predominantly Caucasian Patients with Hidradenitis Suppurativa.

J Invest Dermatol 2020 10 3;140(10):2085-2088.e14. Epub 2020 Mar 3.

Laboratory of Genetic Skin Diseases, INSERM UMR1163 Imagine Institute, Paris, France; Paris University, Paris, France; European Hidradenitis Suppurativa Foundation e.V., Dessau, Germany; Department of Genetics, Necker-Enfants Malades Hospital, AP-HP, Paris, France. Electronic address:

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http://dx.doi.org/10.1016/j.jid.2019.10.025DOI Listing
October 2020

[Twenty years of on-site clinical genetics consultations for people with ASD].

Med Sci (Paris) 2019 Nov 17;35(11):843-851. Epub 2019 Dec 17.

Fédération de Génétique Médicale et Institute Imagine, UMR Inserm 1163, Université Paris-Descartes, Hôpital Necker Enfants-Malades et Fondation Elan Retrouvé, 149 rue de Sèvres, 75015 Paris, France.

Despite advances in neurogenetics of autism spectrum disorders (ASD), many patients fail to be systematically investigated, owing to preconceived ideas, limited access to genetics facilities and inadequacy of consultations to children with behavioural problems. To improve access to services, we reversed the paradigm and delivered on-site genetics consultations to ASD children of Greater Paris day care hospitals and specialized institutions. Since 1998, an ambulatory medical genetics team has been in operation, offering on-site consultations and services to patients and relatives in their usual environment. Because the mobile medical genetics unit operates under the umbrella of a university hospital, service laboratories were shared, including molecular cytogenetics and next generation sequencing (NGS). For the past 20 years, 502 patients from 26 institutions benefited from on-site consultations and genetics services in their usual environment. Less than 1 % of parents declined the offer. Previously undiagnosed genetics conditions were recognized in 71 ASD children, including pathogenic CNV variants (34/388 : 8.8 ; de novo : 19, inherited : 4), Fragile X (4/312 : 1.3 %) and deleterious variants in disease causing genes (33/141 ; 23.4 % : de novo : 23 ; inherited : 10, including 5 X-linked and 5 compound heterozygote mutations). Brain MRI were possible in 347 patients and 42 % were considered abnormal (146/347). All diagnosed patients presented atypical/syndromic ASD with moderate to severe intellectual disability. Thanks to such flexible organisation, a considerable number of missed consultations were tracked and families first benefited from medical genetics services. Owing to constraints imposed by behavioural problems in ASD, we suggest considering on-site genetics services to implement standard of care and counteract the loss of chance to patients and relatives.
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http://dx.doi.org/10.1051/medsci/2019170DOI Listing
November 2019

Identification of an Endoglin Variant Associated With HCV-Related Liver Fibrosis Progression by Next-Generation Sequencing.

Front Genet 2019 4;10:1024. Epub 2019 Nov 4.

Laboratory of Human Genetics of Infectious Diseases, Necker Branch, Inserm U1163, Paris, France.

Despite the astonishing progress in treating chronic hepatitis C virus (HCV) infection with direct-acting antiviral agents, liver fibrosis remains a major health concern in HCV infected patients, in particular due to the treatment cost and insufficient HCV screening in many countries. Only a fraction of patients with chronic HCV infection develop liver fibrosis. While there is evidence that host genetic factors are involved in the development of liver fibrosis, the common variants identified so far, in particular by genome-wide association studies, were found to have limited effects. Here, we conducted an exome association study in 88 highly selected HCV-infected patients with and without fibrosis. A strategy focusing on TGF-β pathway genes revealed an enrichment in rare variants of the endoglin gene () in fibrosis patients. Replication studies in additional cohorts (617 patients) identified one specific variant, Thr5Met, with an overall odds ratio for fibrosis development in carriers of 3.04 (1.39-6.69). Our results suggest that endoglin, a key player in TGF-β signaling, is involved in HCV-related liver fibrogenesis.
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http://dx.doi.org/10.3389/fgene.2019.01024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6844190PMC
November 2019

Impact of on-site clinical genetics consultations on diagnostic rate in children and young adults with autism spectrum disorder.

Mol Autism 2019 7;10:33. Epub 2019 Aug 7.

Fondation Elan Retrouvé, Paris, France.

Background: Neurogenetics investigations and diagnostic yield in patients with autism spectrum disorder (ASD) have significantly improved over the last few years. Yet, many patients still fail to be systematically investigated.

Methods: To improve access to services, an ambulatory team has been established since 1998, delivering on-site clinical genetics consultations and gradually upgrading services to 502 children and young adults with ASD in their standard environment across 26 day-care hospitals and specialized institutions within the Greater Paris region. The evaluation included a clinical genetics consultation, screening for fragile X syndrome, metabolic workup, chromosomal microarray analysis, and, in a proportion of patients, next-generation sequencing of genes reported in ASD and other neurodevelopmental disorders.

Results: Fragile X syndrome and pathogenic copy number variants (CNVs) accounted for the disease in 10% of cases, including 4/312 (1.3%) with fragile X syndrome and 34/388 (8.8%) with pathogenic CNVs (19 de novo and 4 inherited). Importantly, adding high-throughput resequencing of reported intellectual disability/ASD genes to the screening procedure had a major impact on diagnostic yield in the 141 patients examined most recently. Pathogenic or likely pathogenic sequence variants in 27 disease genes were identified in 33/141 patients (23.4%; 23 were de novo and 10 inherited, including five X-linked and five recessive compound heterozygous variants). Diagnosed cases presented atypical and/or syndromic ASD with moderate to severe intellectual disability. The diagnostic yield of fragile X syndrome and array CGH testing combined with next-generation sequencing was significantly higher than fragile X syndrome and array CGH alone ( value 0.009). No inborn errors of metabolism were detected with the metabolic screening.

Conclusion: Based on the diagnostic rate observed in this cohort, we suggest that a stepwise procedure be considered, first screening pathogenic CNVs and a limited number of disease genes in a much larger number of patients, especially those with syndromic ASD and intellectual disability.
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http://dx.doi.org/10.1186/s13229-019-0284-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6686526PMC
June 2020

Pediatric Evans syndrome is associated with a high frequency of potentially damaging variants in immune genes.

Blood 2019 07 2;134(1):9-21. Epub 2019 Apr 2.

Centre de Référence National des Cytopénies Auto-immunes de l'Enfant (CEREVANCE) and.

Evans syndrome (ES) is a rare severe autoimmune disorder characterized by the combination of autoimmune hemolytic anemia and immune thrombocytopenia. In most cases, the underlying cause is unknown. We sought to identify genetic defects in pediatric ES (pES), based on a hypothesis of strong genetic determinism. In a national, prospective cohort of 203 patients with early-onset ES (median [range] age at last follow-up: 16.3 years ([1.2-41.0 years]) initiated in 2004, 80 nonselected consecutive individuals underwent genetic testing. The clinical data were analyzed as a function of the genetic findings. Fifty-two patients (65%) received a genetic diagnosis (the M+ group): 49 carried germline mutations and 3 carried somatic variants. Thirty-two (40%) had pathogenic mutations in 1 of 9 genes known to be involved in primary immunodeficiencies (, , , , , , , , and ), whereas 20 patients (25%) carried probable pathogenic variants in 16 genes that had not previously been reported in the context of autoimmune disease. Lastly, no genetic abnormalities were found in the remaining 28 patients (35%, the M- group). The M+ group displayed more severe disease than the M- group, with a greater frequency of additional immunopathologic manifestations and a greater median number of lines of treatment. Six patients (all from the M+ group) died during the study. In conclusion, pES was potentially genetically determined in at least 65% of cases. Systematic, wide-ranging genetic screening should be offered in pES; the genetic findings have prognostic significance and may guide the choice of a targeted treatment.
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http://dx.doi.org/10.1182/blood-2018-11-887141DOI Listing
July 2019

Copy number variations and founder effect underlying complete IL-10Rβ deficiency in Portuguese kindreds.

PLoS One 2018 26;13(10):e0205826. Epub 2018 Oct 26.

INSERM, UMR1163 and Institut Imagine, Laboratory of Intestinal Immunity, Paris, France.

Mutations in interleukin-10 receptor (IL-10R) genes are one cause of very early-onset inflammatory bowel disease with perianal lesions, which can be cured by hematopoietic stem cell transplantation. Using a functional test, which assesses responsiveness of peripheral monocytes to IL-10, we identified three unrelated Portuguese patients carrying two novel IL-10RB mutations. In the three patients, sequencing of genomic DNA identified the same large deletion of exon 3 which precluded protein expression. This mutation was homozygous in two patients born from consanguineous families and heterozygous in the third patient born from unrelated parents. Microsatellite analysis of the IL10RB genomic region revealed a common haplotype in the three Portuguese families pointing to a founder deletion inherited from a common ancestor 400 years ago. In the third patient, surface expression of IL-10R was normal but signaling in response to IL-10 was impaired. Complementary DNA sequencing and next-generation sequencing of IL10RB locus with custom-made probes revealed a ≈ 6 Kb duplication encompassing the exon 6 which leads to a frameshift mutation and a loss of the TYK2-interacting Box 2 motif. Altogether, we describe two novel copy number variations in IL10RB, one with founder effect and one preserving cell surface expression but abolishing signaling.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0205826PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6203366PMC
April 2019

De novo mutation screening in childhood-onset cerebellar atrophy identifies gain-of-function mutations in the CACNA1G calcium channel gene.

Brain 2018 07;141(7):1998-2013

Paris Descartes - Sorbonne Paris Cité University, Imagine Institute, Paris, France.

Cerebellar atrophy is a key neuroradiological finding usually associated with cerebellar ataxia and cognitive development defect in children. Unlike the adult forms, early onset cerebellar atrophies are classically described as mostly autosomal recessive conditions and the exact contribution of de novo mutations to this phenotype has not been assessed. In contrast, recent studies pinpoint the high prevalence of pathogenic de novo mutations in other developmental disorders such as intellectual disability, autism spectrum disorders and epilepsy. Here, we investigated a cohort of 47 patients with early onset cerebellar atrophy and/or hypoplasia using a custom gene panel as well as whole exome sequencing. De novo mutations were identified in 35% of patients while 27% had mutations inherited in an autosomal recessive manner. Understanding if these de novo events act through a loss or a gain of function effect is critical for treatment considerations. To gain a better insight into the disease mechanisms causing these cerebellar defects, we focused on CACNA1G, a gene not yet associated with the early-onset form. This gene encodes the Cav3.1 subunit of T-type calcium channels highly expressed in Purkinje neurons and deep cerebellar nuclei. We identified four patients with de novo CACNA1G mutations. They all display severe motor and cognitive impairment, cerebellar atrophy as well as variable features such as facial dysmorphisms, digital anomalies, microcephaly and epilepsy. Three subjects share a recurrent c.2881G>A/p.Ala961Thr variant while the fourth patient has the c.4591A>G/p.Met1531Val variant. Both mutations drastically impaired channel inactivation properties with significantly slower kinetics (∼5 times) and negatively shifted potential for half-inactivation (>10 mV). In addition, these two mutations increase neuronal firing in a cerebellar nuclear neuron model and promote a larger window current fully inhibited by TTA-P2, a selective T-type channel blocker. This study highlights the prevalence of de novo mutations in early-onset cerebellar atrophy and demonstrates that A961T and M1531V are gain of function mutations. Moreover, it reveals that aberrant activity of Cav3.1 channels can markedly alter brain development and suggests that this condition could be amenable to treatment.
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http://dx.doi.org/10.1093/brain/awy145DOI Listing
July 2018

Diagnostic Yield of Next-generation Sequencing in Very Early-onset Inflammatory Bowel Diseases: A Multicentre Study.

J Crohns Colitis 2018 08;12(9):1104-1112

GENIUS Group [GENetically ImmUne-mediated enteropathieS] from the European Society for Paediatric Gastroenterology, Hepatology and Nutrition [ESPGHAN].

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.
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http://dx.doi.org/10.1093/ecco-jcc/jjy068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6113703PMC
August 2018

NDUFB8 Mutations Cause Mitochondrial Complex I Deficiency in Individuals with Leigh-like Encephalomyopathy.

Am J Hum Genet 2018 03 8;102(3):460-467. Epub 2018 Feb 8.

Department of Pediatrics, Salzburger Landeskliniken and Paracelsus Medical University Salzburg, 5020 Salzburg, Austria. Electronic address:

Respiratory chain complex I deficiency is the most frequently identified biochemical defect in childhood mitochondrial diseases. Clinical symptoms range from fatal infantile lactic acidosis to Leigh syndrome and other encephalomyopathies or cardiomyopathies. To date, disease-causing variants in genes coding for 27 complex I subunits, including 7 mitochondrial DNA genes, and in 11 genes encoding complex I assembly factors have been reported. Here, we describe rare biallelic variants in NDUFB8 encoding a complex I accessory subunit revealed by whole-exome sequencing in two individuals from two families. Both presented with a progressive course of disease with encephalo(cardio)myopathic features including muscular hypotonia, cardiac hypertrophy, respiratory failure, failure to thrive, and developmental delay. Blood lactate was elevated. Neuroimaging disclosed progressive changes in the basal ganglia and either brain stem or internal capsule. Biochemical analyses showed an isolated decrease in complex I enzymatic activity in muscle and fibroblasts. Complementation studies by expression of wild-type NDUFB8 in cells from affected individuals restored mitochondrial function, confirming NDUFB8 variants as the cause of complex I deficiency. Hereby we establish NDUFB8 as a relevant gene in childhood-onset mitochondrial disease.
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http://dx.doi.org/10.1016/j.ajhg.2018.01.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5985356PMC
March 2018

Mutations in ACTRT1 and its enhancer RNA elements lead to aberrant activation of Hedgehog signaling in inherited and sporadic basal cell carcinomas.

Nat Med 2017 Oct 4;23(10):1226-1233. Epub 2017 Sep 4.

Genoscope (CEA), CNRS UMR 8030, University of Evry, Evry, France.

Basal cell carcinoma (BCC), the most common human cancer, results from aberrant activation of the Hedgehog signaling pathway. Although most cases of BCC are sporadic, some forms are inherited, such as Bazex-Dupré-Christol syndrome (BDCS)-a cancer-prone genodermatosis with an X-linked, dominant inheritance pattern. We have identified mutations in the ACTRT1 gene, which encodes actin-related protein T1 (ARP-T1), in two of the six families with BDCS that were examined in this study. High-throughput sequencing in the four remaining families identified germline mutations in noncoding sequences surrounding ACTRT1. These mutations were located in transcribed sequences encoding enhancer RNAs (eRNAs) and were shown to impair enhancer activity and ACTRT1 expression. ARP-T1 was found to directly bind to the GLI1 promoter, thus inhibiting GLI1 expression, and loss of ARP-T1 led to activation of the Hedgehog pathway in individuals with BDCS. Moreover, exogenous expression of ACTRT1 reduced the in vitro and in vivo proliferation rates of cell lines with aberrant activation of the Hedgehog signaling pathway. In summary, our study identifies a disease mechanism in BCC involving mutations in regulatory noncoding elements and uncovers the tumor-suppressor properties of ACTRT1.
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http://dx.doi.org/10.1038/nm.4368DOI Listing
October 2017

Neutropenia in Patients with Common Variable Immunodeficiency: a Rare Event Associated with Severe Outcome.

J Clin Immunol 2017 Oct 26;37(7):715-726. Epub 2017 Aug 26.

Department of Clinical Immunology and Internal Medicine, Strasbourg University Hospital, National Reference Center for Autoimmune Diseases, Strasbourg, France.

Background: Common variable immunodeficiency (CVID) is characterized by infections and hypogammaglobulinemia. Neutropenia is rare during CVID.

Methods: The French DEFI study enrolled patients with primary hypogammaglobulinemia. Patients with CVID and neutropenia were retrospectively analyzed.

Results: Among 473 patients with CVID, 16 patients displayed neutropenia (lowest count [0-1400]*10/L). Sex ratio (M/F) was 10/6. Five patients died during the follow-up (11 years) with an increased percentage of deaths compared to the whole DEFI group (31.3 vs 3.4%, P < 0.05). Neutropenia was diagnosed for 10 patients before 22 years old. The most frequent symptoms, except infections, were autoimmune cytopenia, i.e., thrombopenia or anemia (11/16). Ten patients were affected with lymphoproliferative diseases. Two patients were in the infection only group and the others belonged to one or several other CVID groups. The median level of IgG was 2.6 g/L [0.35-4.4]. Most patients presented increased numbers of CD21 CD38 B cell, as already described in CVID autoimmune cytopenia group. Neutropenia was considered autoimmune in 11 cases. NGS for 52 genes of interest was performed on 8 patients. No deleterious mutations were found in LRBA, CTLA4, and PIK3. More than one potentially damaging variant in other genes associated with CVID were present in most patients arguing for a multigene process.

Conclusion: Neutropenia is generally associated with another cytopenia and presumably of autoimmune origin during CVID. In the DEFI study, neutropenia is coupled with more severe clinical outcomes. It appears as an "alarm bell" considering patients' presentation and the high rate of deaths. Whole exome sequencing diagnosis should improve management.
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http://dx.doi.org/10.1007/s10875-017-0434-2DOI Listing
October 2017

Compound heterozygosity for severe and hypomorphic mutations cause non-syndromic LHON-like optic neuropathy.

J Med Genet 2017 05 28;54(5):346-356. Epub 2016 Dec 28.

Laboratory of Genetics in Ophthalmology (LGO), INSERM UMR1163, Institute of Genetic Diseases, Imagine, Paris Descartes University, Paris, France.

Background: Non-syndromic hereditary optic neuropathy (HON) has been ascribed to mutations in mitochondrial fusion/fission dynamics genes, nuclear and mitochondrial DNA-encoded respiratory enzyme genes or nuclear genes of poorly known mitochondrial function. However, the disease causing gene remains unknown in many families. The objective of the present study was to identify the molecular cause of non-syndromic LHON-like disease in siblings born to non-consanguineous parents of French origin.

Methods: We used a combination of genetic analysis (gene mapping and whole-exome sequencing) in a multiplex family of non-syndromic HON and of functional analyses in patient-derived cultured skin fibroblasts and the yeast .

Results: We identified compound heterozygote disease-causing mutations (p.Tyr53Cys; p.Tyr308Cys). Studies using patient-derived cultured skin fibroblasts revealed mildly decreased NDUFS2 and complex I abundance but apparently normal respiratory chain activity. In the yeast ortholog , the mutations resulted in absence of complex I and moderate reduction in nicotinamide adenine dinucleotide-ubiquinone oxidoreductase activity, respectively.

Conclusions: Biallelism for mutations causing severe complex I deficiency has been previously reported to cause Leigh syndrome with optic neuropathy. Our results are consistent with the view that compound heterozygosity for severe and hypomorphic mutations can cause non-syndromic HON. This observation suggests a direct correlation between the severity of mutations and that of the disease and further support that there exist a genetic overlap between non-syndromic and syndromic HON due to defective mitochondrial function.
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http://dx.doi.org/10.1136/jmedgenet-2016-104212DOI Listing
May 2017

Exome sequencing links corticospinal motor neuron disease to common neurodegenerative disorders.

Science 2014 Jan;343(6170):506-511

Department of Pediatrics and Neonatology, Saudi German Hospital, Post Office Box 84348, Riyadh, Kingdom of Saudi Arabia.

Hereditary spastic paraplegias (HSPs) are neurodegenerative motor neuron diseases characterized by progressive age-dependent loss of corticospinal motor tract function. Although the genetic basis is partly understood, only a fraction of cases can receive a genetic diagnosis, and a global view of HSP is lacking. By using whole-exome sequencing in combination with network analysis, we identified 18 previously unknown putative HSP genes and validated nearly all of these genes functionally or genetically. The pathways highlighted by these mutations link HSP to cellular transport, nucleotide metabolism, and synapse and axon development. Network analysis revealed a host of further candidate genes, of which three were mutated in our cohort. Our analysis links HSP to other neurodegenerative disorders and can facilitate gene discovery and mechanistic understanding of disease.
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http://dx.doi.org/10.1126/science.1247363DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4157572PMC
January 2014

TMEM126A is a mitochondrial located mRNA (MLR) protein of the mitochondrial inner membrane.

Biochim Biophys Acta 2013 Jun 13;1830(6):3719-33. Epub 2013 Mar 13.

Départment de Génétique, Fondation Imagine, Université Paris Descartes-Sorbonne Paris Cité, Paris, France.

Background: Hereditary optic neuropathies (HONs) are a heterogeneous group of disorders that affect retinal ganglion cells (RGCs) and axons that form the optic nerve. Leber's Hereditary Optic Neuropathy and the autosomal dominant optic atrophy related to OPA1 mutations are the most common forms. Nonsyndromic autosomal recessive optic neuropathies are rare and their existence has been long debated. We recently identified the first gene responsible for these conditions, TMEM126A. This gene is highly expressed in retinal cellular compartments enriched in mitochondria and supposed to encode a mitochondrial transmembrane protein of unknown function.

Methods: A specific polyclonal antibody targeting the TMEM126A protein has been generated. Quantitative fluorescent in situ hybridization, cellular fractionation, mitochondrial membrane association study, mitochondrial sub compartmentalization analysis by both proteolysis assays and transmission electron microscopy, and expression analysis of truncated TMEM126A constructs by immunofluorescence confocal microscopy were carried out.

Results: TMEM126A mRNAs are strongly enriched in the vicinity of mitochondria and encode an inner mitochondrial membrane associated cristae protein. Moreover, the second transmembrane domain of TMEM126A is required for its mitochondrial localization.

Conclusions: TMEM126A is a mitochondrial located mRNA (MLR) that may be translated in the mitochondrial surface and the protein is subsequently imported to the inner membrane. These data constitute the first step toward a better understanding of the mechanism of action of TMEM126A in RGCs and support the importance of mitochondrial dysfunction in the pathogenesis of HON.

General Significance: Local translation of nuclearly encoded mitochondrial mRNAs might be a mechanism for rapid onsite supply of mitochondrial membrane proteins.
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http://dx.doi.org/10.1016/j.bbagen.2013.02.025DOI Listing
June 2013

AON-mediated Exon Skipping Restores Ciliation in Fibroblasts Harboring the Common Leber Congenital Amaurosis CEP290 Mutation.

Mol Ther Nucleic Acids 2012 Jun 26;1:e29. Epub 2012 Jun 26.

1] Genethon, Evry, France [2] CNRS UMR 8151-Inserm U1022, Université Paris Descarte-Sorbonne Paris Cité, Chimie-Paristech, Paris, France.

Leber congenital amaurosis (LCA) is a severe hereditary retinal dystrophy responsible for congenital or early-onset blindness. The most common disease-causing mutation (>10%) is located deep in intron 26 of the CEP290 gene (c.2991+1655A>G). It creates a strong splice donor site that leads to insertion of a cryptic exon encoding a premature stop codon. In the present study, we show that the use of antisense oligonucleotides (AONs) allow an efficient skipping of the mutant cryptic exon and the restoration of ciliation in fibroblasts of affected patients. These data support the feasibility of an AON-mediated exon skipping strategy to correct the aberrant splicing.
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http://dx.doi.org/10.1038/mtna.2012.21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3390222PMC
June 2012

ALDH1A3 mutations cause recessive anophthalmia and microphthalmia.

Am J Hum Genet 2013 Feb 9;92(2):265-70. Epub 2013 Jan 9.

INSERM U781 & Department of Genetics, Paris Descartes University, Paris, France.

Anophthalmia and microphthalmia (A/M) are early-eye-development anomalies resulting in absent or small ocular globes, respectively. A/M anomalies occur in syndromic or nonsyndromic forms. They are genetically heterogeneous, some mutations in some genes being responsible for both anophthalmia and microphthalmia. Using a combination of homozygosity mapping, exome sequencing, and Sanger sequencing, we identified homozygosity for one splice-site and two missense mutations in the gene encoding the A3 isoform of the aldehyde dehydrogenase 1 (ALDH1A3) in three consanguineous families segregating A/M with occasional orbital cystic, neurological, and cardiac anomalies. ALDH1A3 is a key enzyme in the formation of a retinoic acid gradient along the dorso-ventral axis during early eye development. Transitory expression of mutant ALDH1A3 open reading frames showed that both missense mutations reduce the accumulation of the enzyme, potentially leading to altered retinoic acid synthesis. Although the role of retinoic acid signaling in eye development is well established, our findings provide genetic evidence of a direct link between retinoic-acid-synthesis dysfunction and early-eye-development anomalies in humans.
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http://dx.doi.org/10.1016/j.ajhg.2012.12.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3567280PMC
February 2013

Mutations in NMNAT1 cause Leber congenital amaurosis with early-onset severe macular and optic atrophy.

Nat Genet 2012 Sep 29;44(9):975-7. Epub 2012 Jul 29.

Department of Genetics, Institut National de la Santé et de la Recherche Médicale U, Université Paris Descartes-Sorbonne Paris Cité, France.

In addition to its activity in nicotinamide adenine dinucleotide (NAD(+)) synthesis, the nuclear nicotinamide mononucleotide adenyltransferase NMNAT1 acts as a chaperone that protects against neuronal activity-induced degeneration. Here we report that compound heterozygous and homozygous NMNAT1 mutations cause severe neonatal neurodegeneration of the central retina and early-onset optic atrophy in 22 unrelated individuals. Their clinical presentation is consistent with Leber congenital amaurosis and suggests that the mutations affect neuroprotection of photoreceptor cells.
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http://dx.doi.org/10.1038/ng.2357DOI Listing
September 2012

Mainzer-Saldino syndrome is a ciliopathy caused by IFT140 mutations.

Am J Hum Genet 2012 May 12;90(5):864-70. Epub 2012 Apr 12.

INSERM U781 & Department of Genetics, Paris Descartes University, Paris, France.

Mainzer-Saldino syndrome (MSS) is a rare disorder characterized by phalangeal cone-shaped epiphyses, chronic renal failure, and early-onset, severe retinal dystrophy. Through a combination of ciliome resequencing and Sanger sequencing, we identified IFT140 mutations in six MSS families and in a family with the clinically overlapping Jeune syndrome. IFT140 is one of the six currently known components of the intraflagellar transport complex A (IFT-A) that regulates retrograde protein transport in ciliated cells. Ciliary abundance and localization of anterograde IFTs were altered in fibroblasts of affected individuals, a result that supports the pivotal role of IFT140 in proper development and function of ciliated cells.
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http://dx.doi.org/10.1016/j.ajhg.2012.03.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3376548PMC
May 2012

Mutation in a primate-conserved retrotransposon reveals a noncoding RNA as a mediator of infantile encephalopathy.

Proc Natl Acad Sci U S A 2012 Mar 12;109(13):4980-5. Epub 2012 Mar 12.

Département de Génétique and Groupe de Recherche Immunopathologies et Maladies Infectieuses, Université de La Réunion, Centre Hospitalier Régional de La Réunion, 97405 Saint-Denis, La Réunion, France.

The human genome is densely populated with transposons and transposon-like repetitive elements. Although the impact of these transposons and elements on human genome evolution is recognized, the significance of subtle variations in their sequence remains mostly unexplored. Here we report homozygosity mapping of an infantile neurodegenerative disease locus in a genetic isolate. Complete DNA sequencing of the 400-kb linkage locus revealed a point mutation in a primate-specific retrotransposon that was transcribed as part of a unique noncoding RNA, which was expressed in the brain. In vitro knockdown of this RNA increased neuronal apoptosis, consistent with the inappropriate dosage of this RNA in vivo and with the phenotype. Moreover, structural analysis of the sequence revealed a small RNA-like hairpin that was consistent with the putative gain of a functional site when mutated. We show here that a mutation in a unique transposable element-containing RNA is associated with lethal encephalopathy, and we suggest that RNAs that harbor evolutionarily recent repetitive elements may play important roles in human brain development.
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http://dx.doi.org/10.1073/pnas.1111596109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3323976PMC
March 2012

Intellectual disability associated with retinal dystrophy in the Xp11.3 deletion syndrome: ZNF674 on trial. Guilty or innocent?

Eur J Hum Genet 2012 Mar 30;20(3):352-6. Epub 2011 Nov 30.

INSERM U781 - Department of Genetics/Fondation IMAGINE and Paris Descartes University, CHU Necker Enfants Malades, Paris, France.

X-linked retinal dystrophies (XLRD) are listed among the most severe RD owing to their early onset, leading to significant visual loss before the age of 30. One-third of XLRD are accounted for by RP2 mutations at the Xp11.23 locus. Deletions of ca. 1.2 Mb in the Xp11.3-p11.23 region have been previously reported in two independent families segregating XLRD with intellectual disability (ID). Although the RD was ascribed to the deletion of RP2, the ID was suggested to be accounted for by the loss of ZNF674, which mutations were independently reported to account for isolated XLID. Here, we report deletions in the Xp11.3-p11.23 region responsible for the loss of ZNF674 in two unrelated families segregating XLRD, but no ID, identified by chromosomal microarray analysis. These findings question the responsibility of ZNF674 deletions in ID associated with X-linked retinal dystrophy.
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http://dx.doi.org/10.1038/ejhg.2011.217DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3283176PMC
March 2012

Nuclear outsourcing of RNA interference components to human mitochondria.

PLoS One 2011 13;6(6):e20746. Epub 2011 Jun 13.

INSERM U781 Hôpital Necker-Enfants Malades, Paris, France.

MicroRNAs (miRNAs) are small non-coding RNAs that associate with Argonaute proteins to regulate gene expression at the post-transcriptional level in the cytoplasm. However, recent studies have reported that some miRNAs localize to and function in other cellular compartments. Mitochondria harbour their own genetic system that may be a potential site for miRNA mediated post-transcriptional regulation. We aimed at investigating whether nuclear-encoded miRNAs can localize to and function in human mitochondria. To enable identification of mitochondrial-enriched miRNAs, we profiled the mitochondrial and cytosolic RNA fractions from the same HeLa cells by miRNA microarray analysis. Mitochondria were purified using a combination of cell fractionation and immunoisolation, and assessed for the lack of protein and RNA contaminants. We found 57 miRNAs differentially expressed in HeLa mitochondria and cytosol. Of these 57, a signature of 13 nuclear-encoded miRNAs was reproducibly enriched in mitochondrial RNA and validated by RT-PCR for hsa-miR-494, hsa-miR-1275 and hsa-miR-1974. The significance of their mitochondrial localization was investigated by characterizing their genomic context, cross-species conservation and instrinsic features such as their size and thermodynamic parameters. Interestingly, the specificities of mitochondrial versus cytosolic miRNAs were underlined by significantly different structural and thermodynamic parameters. Computational targeting analysis of most mitochondrial miRNAs revealed not only nuclear but also mitochondrial-encoded targets. The functional relevance of miRNAs in mitochondria was supported by the finding of Argonaute 2 localization to mitochondria revealed by immunoblotting and confocal microscopy, and further validated by the co-immunoprecipitation of the mitochondrial transcript COX3. This study provides the first comprehensive view of the localization of RNA interference components to the mitochondria. Our data outline the molecular bases for a novel layer of crosstalk between nucleus and mitochondria through a specific subset of human miRNAs that we termed 'mitomiRs'.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0020746PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3113838PMC
November 2011

Spectrum of SPATA7 mutations in Leber congenital amaurosis and delineation of the associated phenotype.

Hum Mutat 2010 Mar;31(3):E1241-50

Unité de Recherches Génétique et Epigénétique des Maladies Métaboliques, Neurosensorielles et du Développement, INSERM U781 and Université Paris Descartes, CHU Necker Enfants Malades, Paris, France.

Leber congenital amaurosis (LCA) is the earliest and most severe retinal degeneration. It may present as a congenital stationary cone-rod dystrophy (LCA type I) or a progressive yet severe rod-cone dystrophy (LCA type II). Twelve LCA genes have been identified, three of which account for Type I and nine for LCA type II. All proteins encoded by these genes but two are preferentially expressed in the retina and are responsible for non-syndromic LCA only. By contrast LCA5 and CEP290 are widely expressed and mutations in this latter result in a variety of phenotypes from non-syndromic retinal degeneration to pleiotropic disorders including senior-Loken (SNLS) and Joubert syndromes (JBTS). Recently, mutations in the widely expressed gene SPATA7 were reported to cause LCA or juvenile retinitis pigmentosa. The purpose of this study was i) to determine the level of expression of two major alternative SPATA7 transcripts in a large range of tissues and ii) to assess the involvement of this novel gene in a large cohort of unrelated patients affected with LCA (n = 134). Here, we report high SPATA7expression levels in retina, brain and testis with differential expression of the two transcripts. SPATA7 mutations were identified in few families segregating non-syndromic LCA (n = 4/134). Six different mutations were identified, four of which are novel; All affected both SPATA7 transcripts. The clinical evaluation of patients suggested that SPATA7 mutations account for the rod-cone dystrophy type of the disease.
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http://dx.doi.org/10.1002/humu.21203DOI Listing
March 2010

TMEM126A, encoding a mitochondrial protein, is mutated in autosomal-recessive nonsyndromic optic atrophy.

Am J Hum Genet 2009 Apr 26;84(4):493-8. Epub 2009 Mar 26.

Département de Génétique, Université Paris Descartes, Unité INSERM U781, Hôpital Necker-Enfants Malades, 75015 Paris, France.

Nonsyndromic autosomal-recessive optic neuropathies are rare conditions of unknown genetic and molecular origin. Using an approach of whole-genome homozygosity mapping and positional cloning, we have identified the first gene, to our knowledge, responsible for this condition, TMEM126A, in a large multiplex inbred Algerian family and subsequently in three other families originating from the Maghreb. TMEM126A is conserved in higher eukaryotes and encodes a transmembrane mitochondrial protein of unknown function, supporting the view that mitochondrial dysfunction may be a hallmark of inherited optic neuropathies including isolated autosomal-recessive forms.
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http://dx.doi.org/10.1016/j.ajhg.2009.03.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2667974PMC
April 2009

Identification of the SPG15 gene, encoding spastizin, as a frequent cause of complicated autosomal-recessive spastic paraplegia, including Kjellin syndrome.

Am J Hum Genet 2008 Apr;82(4):992-1002

Institut National de la Santé et de la Recherche Médicale (INSERM), Unité Mixte de Recherche (UMR) S679, Neurologie et Thérapeutique Expérimentale, Paris, F-75013 France.

Hereditary spastic paraplegias (HSPs) are genetically and phenotypically heterogeneous disorders. Both "uncomplicated" and "complicated" forms have been described with various modes of inheritance. Sixteen loci for autosomal-recessive "complicated" HSP have been mapped. The SPG15 locus was first reported to account for a rare form of spastic paraplegia variably associated with mental impairment, pigmented maculopathy, dysarthria, cerebellar signs, and distal amyotrophy, sometimes designated as Kjellin syndrome. Here, we report the refinement of SPG15 to a 2.64 Mb genetic interval on chromosome 14q23.3-q24.2 and the identification of ZFYVE26, which encodes a zinc-finger protein with a FYVE domain that we named spastizin, as the cause of SPG15. Six different truncating mutations were found to segregate with the disease in eight families with a phenotype that included variable clinical features of Kjellin syndrome. ZFYVE26 mRNA was widely distributed in human tissues, as well as in rat embryos, suggesting a possible role of this gene during embryonic development. In the adult rodent brain, its expression profile closely resembled that of SPG11, another gene responsible for complicated HSP. In cultured cells, spastizin colocalized partially with markers of endoplasmic reticulum and endosomes, suggesting a role in intracellular trafficking.
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http://dx.doi.org/10.1016/j.ajhg.2008.03.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2427184PMC
April 2008

Mutations in LCA5 are an uncommon cause of Leber congenital amaurosis (LCA) type II.

Hum Mutat 2007 Dec;28(12):1245

INSERM U781 et Service de Génétique, Hôpital des Enfants Malades, 149 rue de Sèvres, 75743 Paris Cedex 15, France.

Leber congenital amaurosis (LCA) is the earliest and most severe form of inherited retinal dystrophy responsible for blindness or severe visual impairment at birth or within the first months of life. Up to date, ten LCA genes have been identified. Three of them account for ca. 43% of families and are responsible for a congenital severe stationary cone-rod dystrophy (Type I, 60% of LCA) while the seven remaining genes account for 32% of patients and are responsible for a progressive yet severe rod-cone dystrophy (Type II, 40% of LCA ). Recently, mutations in LCA5, encoding the ciliary protein lebercilin, were reported to be a rare cause of leber congenital amaurosis. The purpose of this study was to evaluate the involvement of this novel gene and to look for genotype-phenotype correlations. Here we report the identification of three novel LCA5 mutations (3/3 homozygous) in three families confirming the modest implication of this gene in our series (3/179; 1.7%). Besides, we suggest that the phenotype of these patients affected with a particularly severe form of LCA type II may represent a continuum with LCA type I.
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http://dx.doi.org/10.1002/humu.9513DOI Listing
December 2007
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