Imprinting aberrations of SNRPN, ZAC1 and INPP5F genes involved in the pathogenesis of congenital heart disease with extracardiac malformations.

Congenital heart disease (CHD) with extracardiac malformations (EM) is the most common multiple malformation, resulting from the interaction between genetic abnormalities and environmental factors. Most studies have attributed the causes of CHD with EM to chromosomal abnormalities. However, multi-system dysplasia is usually caused by both genetic mutations and epigenetic dysregulation. The epigenetic mechanisms underlying the pathogenesis of CHD with EM remain unclear. In this study, we investigated the mechanisms of imprinting alterations, including those of the Small nuclear ribonucleoprotein polypeptide N (SNRPN), PLAG1 like zinc finger 1 (ZAC1) and inositol polyphosphate-5-phosphatase F (INPP5F) genes, in the pathogenesis of CHD with EM. The methylation levels ofSNRPN,ZAC1,andINPP5Fgenes were analysed by the MassARRAY platform in 24 children with CHD with EM and 20 healthy controls. The expression levels of these genes were detected by real-time polymerase chain reaction (PCR). The correlation between methylation regulation and gene expression was confirmed using 5-azacytidine (5-Aza) treated cells. The methylation levels ofSNRPNandZAC1genes were significantly increased in CHD with EM, while that ofINPP5Fwas decreased. The methylation alterations of these genes were negatively correlated with expression. Risk analysis showed that abnormal hypermethylation ofSNRPNandZAC1resulted in 5.545 and 7.438 times higher risks of CHD with EM, respectively, and the abnormal hypomethylation ofINPP5Fwas 8.38 times higher than that of the control group. We concluded that abnormally high methylation levels ofSNRPNandZAC1and decreased levels ofINPP5Fimply an increased risk of CHD with EM by altering their gene functions. This study provides evidence of imprinted regulation in the pathogenesis of multiple malformations.