Ventricular septal defects (VSD) are the most common congenital heart defects (CHD). Studies have documented that ISL1 has a crucial impact on cardiac growth, but the role of variants in the ISL1 gene promoter in patients with VSD has not been explored. In 400 subjects (200 isolated and sporadic VSD patients: 200 healthy controls), we investigated the ISL1 gene promoter variant and performed cellular functional experiments by using the dual-luciferase reporter assay to verify the impact on gene expression. In the ISL1 promoter, 5 variants were found only in VSD patients by sequencing. Cellular functional experiments demonstrated that three variants decreased the transcriptional activity of the ISL1 promoter (P < 0.05). Further analysis with the online JASPAR database demonstrated that a cluster of putative binding sites for transcription factors may be altered by these variants, possibly resulting in change of ISL1 protein expression and VSD formation. Our study has for the first time identified novel variants in the ISL1 gene promoter region in the Han Chinese patients with isolated and sporadic VSD. Additionally, the cellular functional experiments, electrophoretic mobility shift assay, and bioinformatic analysis have demonstrated that these variants significantly alter the expression of the ISL1 gene and affect the binding of transcription factors, likely resulting in VSD. Therefore, this study may provide new insights into the role of the gene promoter region for a better understanding of genetic basis of the formation of CHD and may promote further investigations on mechanism of the formation of CHD.

中文翻译：

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孤立室间隔缺损中 ISL1 基因启动子和细胞功能变体的鉴定

室间隔缺损 (VSD) 是最常见的先天性心脏病 (CHD)。研究表明，ISL1 对心脏生长具有至关重要的影响，但尚未探索 ISL1 基因启动子变异在 VSD 患者中的作用。在 400 名受试者（200 名孤立和散发性 VSD 患者：200 名健康对照者）中，我们研究了 ISL1 基因启动子变异并通过使用双荧光素酶报告基因检测进行了细胞功能实验，以验证对基因表达的影响。在 ISL1 启动子中，通过测序仅在 VSD 患者中发现了 5 个变异。细胞功能实验表明，三种变体降低了 ISL1 启动子的转录活性（P < 0.05）。对在线 JASPAR 数据库的进一步分析表明，这些变异可能会改变一组假定的转录因子结合位点，这可能导致 ISL1 蛋白表达和 VSD 形成的变化。我们的研究首次在患有孤立性和散发性 VSD 的汉族患者中发现了 ISL1 基因启动子区域的新变异。此外，细胞功能实验、电泳迁移率变化分析和生物信息学分析表明，这些变异显着改变了 ISL1 基因的表达并影响转录因子的结合，可能导致 VSD。所以，