LMNA Missense Mutation Causes Nonsense-mediated mRNA Decay and Severe Dilated Cardiomyopathy.

Background - LMNA is a known causative gene of dilated cardiomyopathy and familial conduction disturbance. Nonsense-mediated mRNA decay (NMD), normally caused by nonsense mutations, is a safeguard process to protect cells from deleterious effects of inappropriate proteins from mutated genes. NMD induced by non-stop codon mutations is rare. We investigated the effect of an LMNA missense mutation identified in two families affected by cardiac laminopathy. Methods - Genomic DNA and total RNA were isolated from patients' peripheral blood lymphocytes or cardiac tissue. LMNA-coding exons were screened by direct sequencing. Complementary DNAs (cDNAs) were generated by reverse transcription PCR from total RNA. Quantitative PCR (qPCR) was performed to quantify the LMNA cDNA amount by using specific primers for lamins A and C. A minigene splicing reporter experiment was performed to assess the effect of detected variants on RNA splicing. The protein expressions of both isoforms were analyzed by western blotting. Results - We detected a missense variant c.936 G>C (p. Q312H) at the end of exon 5 of LMNA by genomic DNA sequencing in two unrelated families affected by dilated cardiomyopathy and cardiac conduction disturbance. This variant was previously reported in a French family suffering from muscular dystrophy and cardiac conduction disturbance. Sequencing of cDNA demonstrated that the mutated allele was absent. By qPCR assay, we confirmed a 90% reduction in LMNA cDNA. The minigene splicing reporter assay demonstrated a splicing error by the variant. Western blot analysis revealed that lamin A and C expressions were reduced far more than 50%. Conclusions - We report an LMNA missense mutation found in two families, which disrupted a normal splicing site, led to NMD, and resulted in severe cardiac laminopathy.