Background: Despite the accelerated discovery of genes associated with syndromic traits, the majority of families affected by such conditions remain undiagnosed. Here, we carried out an effort to elucidate the underlying genetic defect associated with central nervous system (CNS), cardiac, renal, and digit abnormalities in two unrelated consanguineous families.
Methods and Results: We employed whole exome sequencing to identify homozygous truncating mutations in TMEM260, a locus predicted to encode numerous splice isoforms. Systematic expression analyses across tissues and developmental stages validated two such isoforms, which differ in the utilization of an internal exon. The mutations in both families map uniquely to the long isoform, raising the possibility of an isoform-specific disorder. Consistent with this notion, RT-PCR of lymphocyte cell lines from one of the families showed reduced levels of only the long isoform, which could be ameliorated by emetine, suggesting that the mutation induces nonsense-mediated decay. Subsequent in vivo testing supported this hypothesis. First, either transient suppression or CRISPR/Cas9 genome editing of zebrafish tmem260 recapitulated key neurological phenotypes. Second, co-injection of morphants with the long human TMEM260 mRNA rescued CNS pathology, whereas the short isoform was significantly less efficient. Finally, immunocytochemical and biochemical studies showed preferential enrichment of the long TMEM260 isoform to the plasma membrane.
Conclusion: Reduced functionality of TMEM260 and attenuation of the membrane-associated functions of this protein are principal drivers of pathology, producing CNS, cardiac and renal malformations. These observations contribute to an appreciation of the roles of splice isoforms in genetic disorders.
