Anomalies d'épissage dans les troubles du rythme et cardiomyopathies d'origine génétique (travail effectué dans le laboratoire de Cardiogénétique Moléculaire [du] Professeur Rousson, Centre de Biologie et Pathologie Est [des] Hospices Civils de Lyon)

LYON1-BU Santé (693882101) / SudocSudocFranceF

Combined Use of In Silico and In Vitro Splicing Assays for Interpretation of Genomic Variants of Unknown Significance in Cardiomyopathies and Channelopathies

The identification of molecular anomalies in patients with inherited arrhythmias or cardiomyopathies is a multi challenge due to: i) the number of genes involved; ii) the number of polymorphisms and the fact that most mutations are private; and iii) the variable degree of penetrance which complicates family segregation study. Consequently, a number of unclassified variants (UV) are found in patients’ DNA and some (outside the canonical GT/AG) may affect splicing. Mutational screening on the most prevalent genes involved in arrythmias syndromes or in cardiomyopathies was performed on a cohort made up of 740 unrelated French index probands. To identify splice variants among the identified UVs, a combination of available in silico and in vitro tools were used since transcript is not available. Using this approach, 10 previously identified UVs were reclassified as disease-causing mutations and, more precisely, as haploinsufficiency mutations rather than dominant-negative mutations. Most of them (7 of 10) were observed in MYBPC3. Our study highlighted the importance of the combined use of in silico and in vitro splicing assays to improve the prediction of the functional impact of identified genetic variants. The primary challenge now, with new sequencing technologies, is to distinguish between background polymorphisms and pathogenic mutations. Since splice site mutations can account for almost 50% of disease-causing mutations, recognizing them from among other variations is essential

POLG exon 22 skipping induced by different mechanisms in two unrelated cases of Alpers syndrome.

International audienceThe POLG genes were sequenced in two unrelated patients presenting with Alpers syndrome. The novel c.3626_3629dupGATA and the c.3643+2T>C alleles were associated in trans with p.A467T and p.[W748S;E1143G], respectively. POLG transcripts from skin fibroblasts showed complete exon 22 skipping for patient 2, but surprisingly partial exon 22 skipping from the c.3626_3629dupGATA for patient 1. The creation of a putative exonic splicing silencer could be responsible for the splicing anomaly observed in patient 1. Both c.3643+2T>C and c.3626_3629dupGATA create a premature termination codon and a low polymerase γ activity in skin fibroblasts is responsible for the severe phenotype in these patients

Prevalence and spectrum of mutations in a cohort of 192 unrelated patients with hypertrophic cardiomyopathy.

International audienceHypertrophic Cardiomyopathy (HCM), a common and clinically heterogeneous disease characterized by unexplained ventricular myocardial hypertrophy and a high risk of sudden cardiac death, is mostly caused by mutations in sarcomeric genes but modifiers genes may also modulate the phenotypic expression of HCM mutations. The aim of the current study was to report the frequency of single and multiple gene mutations in a large French cohort of HCM patients and to evaluate the influence of polymorphisms previously suggested to be potential disease modifiers in this myocardial pathology. We report the molecular screening of 192 unrelated HCM patients using denaturing high-performance liquid chromatography/sequencing analysis of the MYBPC3, MYH7, TNNT2 and TNNI3 genes. Genotyping of 6 gene polymorphisms previously reported as putative HCM modifiers (5 RAAS polymorphisms and TNF-α -308 G/A) was also performed. Seventy-five mutations were identified in 92 index patients (48%); 32 were novel. MYBPC3 mutations (25%) represent the most prevalent cause of inherited HCM whereas MYH7 mutations (12%) rank second in the pathogenesis. The onset age was older in patients carrying MYBPC3 mutations than in those with MYH7 mutations. The MYBPC3 IVS20-2A>G splice mutation was identified in 7% of our HCM population. Multiple gene mutations were identified in 9 probands (5%), highlighting the importance of screening other HCM-causing genes even after a first mutation has been identified, particularly in young patients with a severe phenotype. No single or cumulative genetic modifier effect could be evidenced in this HCM cohort