A Common Indel Polymorphism Of The Desmoglein-2 (DSG2) Is Associated With Sudden Cardiac Death In Chinese Populations

Objective:To analyze the connection between the indel polymorphism rs397729601 in the 3' UTR of DSG2 and SCD in a Chinese population,and investigate the underlined mechanisms.Methods:(1)polymorphism screening:we used a candidate gene approach to choose a 2-base pair insertion/deletion(indel)polymorphism(rs397729601)as candidate.(2)case-control study:isolated the genomic DNA from 124 SCD cases and 656 healthy controls,then,PCR combined with Native-PAGE were used to genotyping,the associations between the rs397729601 and the risk of SCD were analyzed using logistic regression,adjusted by sex and age.(3)genotype-phenotype analysis:qPCR analysis was applied to analyze the associations between the relative expression levels of DSG2 mRNA and different genotypic groups.(4)MicroRNA prediction:Miranda was used to predict miRNA which bind with DSG2 3'UTR and influence transcriptional activity.(5)Construction of vector and Dual-luciferase reporter assay:constructed reporter plasmid vector containing insertion or deletion allele of rs397729601(pGL3-control-DSG2-WT/MT)and transfected into 293T cell lines.we investigate the influence of miR-933-3p on 3'UTR of DSG2 through Dual-luciferase reporter assay.Results:(1)The genotyping results showed that the genotype frequencies of the insertion/deletion(indel)polymorphism(rs397729601)in the 3' UTR of DSG2 in the control group were consistent with HWE(P>0.05).It is shown by logistic regression analysis that the risk of SCD has been significantly increased by the deletion allele of rs397729601(OR:1.53,95%CI:1.14-2.04,P=2.97×10-3).(2)Results of qPCR demonstrated that the relative mRNA expression level of DSG2 in samples with ins/del or del/del genotype was 2.69 fold higher than that with ins/ins genotype(P<0.001).(3)Bioinformatic results suggested that DSG2 expression could be regulated by rs397729601 which interrupted the binding of miR-933-3p with DSG2.(4)Results of Dual-luciferase reporter assay indicated that the WT groups drove more decreased reporter expression compared with MT groups in 293T cells with mimic controls;after adding the exogenous miR-933-3p,there was no effective change in MT group,contrasted with mimic control;a significant difference(P<0.001)was found WT groups between miR-933-3p and mimic control group.Conclusions:(1)The rs397729601 polymorphism in the DSG2 gene is associated with SCD risk in Chinese populations.(2)The 2-bp indel polymorphism(rs397729601)within human DSG2 3'UTR may functionally regulate the expression of DSG2 through miR-933-3p and thereby affect development of SCD.(3)The indel polymorphism(rs397729601)may become a useful marker for SCD molecular diagnosis as well as genetic counseling.