First identification of nonsense mutations in autosomal dominant focal segmental glomerulosclerosis.

Recently, a novel heterozygous missense mutation c.T1421G (p. L474R) in the gene encoding podocalyxin was identified in an autosomal dominant focal segmental glomerulosclerosis (AD-FSGS) pedigree. However, this mutation appeared not to impair podocalyxin function, and it is necessary to identify new mutations and determine their causative role for FSGS. In the present study, we report the identification of a heterozygous nonsense mutation (c.C976T; p. Arg326X) in a Chinese pedigree featured by proteinuria and renal insufficiency with AD inheritance by whole exome sequencing (WES). Total mRNA and PODXL protein abundance were decreased in available peripheral blood cell samples of two affected patients undergoing hemodialysis, compared with those in healthy controls and hemodialysis controls without mutation. We identified another novel heterozygous nonsense mutation (c.C1133G; p.Ser378X) in a British-Indian pedigree of AD-FSGS by WES. study showed that, human embryonic kidney 293T cells transfected with the pEGFP-PODXL-Arg326X or pEGFP-PODXL-Ser378X plasmid expressed significantly lower mRNA and PODXL protein compared with cells transfected with the wild-type plasmid. Blocking nonsense-mediated mRNA decay (NMD) significantly restored the amount of mutant mRNA and PODXL proteins, which indicated that the pathogenic effect of nonsense mutations is likely due to NMD, resulting in podocalyxin deficiency. Functional consequences caused by the nonsense mutations were inferred by siRNA knockdown in cultured podocytes and podocalyxin down-regulation by siRNA resulted in decreased RhoA and ezrin activities, cell migration and stress fiber formation. Our results provided new data implicating heterozygous nonsense mutations in the development of FSGS.