[Synonymous variants of the ATP7B gene may cause abnormal splicing of mRNA by affecting the exonic splicing enhancers].

OBJECTIVE:To explore the effect of rare synonymous variants of the ATP7B gene on the splicing of its precursor mRNA. METHODS:A total of 248 rare synonymous variants with allelic frequency of <0.005 were retrieved from the ExAc database. Human Splicing Finder (HSF) was used to predict their effect on the splicing of precursor mRNA. And ESE Finder 3.0 was used to predict the effect of such variants on the binding ability of SR protein family. Rare synonymous variants affecting the binding of two or more SR proteins were selected and verified with an in vitro mini gene splicing report system. RESULTS:HSF analysis indicated that 136 of the 248 rare synonymous variants may destroy the exonic splicing enhancer (ESE) motif. Analysis using ESE Finder 3.0 indicated that 19 of them may affect the binding of two or more SR proteins at the same time. In vitro mini gene experiment confirmed that the c.1620C>T (p.L540L) and c.3888C>T (p.A1296A) variants could lead to abnormal splicing of the corresponding exons, resulting in complete skipping of exon 4 and 25% increase in the skipping of exon 18, respectively. CONCLUSION:Synonymous variants may affect the splicing of precursor mRNA in various ways, particularly the destruction of ESE motif. This study confirmed that the c.1620C>T (p.L540L) and c.3888C>T (p.A1296A) variants can affect the mRNA splicing of the ATP7B gene, resulting in skipping of corresponding exons, which may provide a basis for genetic diagnosis and consultation of carriers.

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