Coronary artery disease genes SMAD3 and TCF21 promote opposing interactive genetic programs that regulate smooth muscle cell differentiation and disease risk

Although numerous genetic loci have been associated with coronary artery disease (CAD) with genome wide association studies, efforts are needed to identify the causal genes in these loci and link them into fundamental signaling pathways. Recent studies have investigated the disease mechanism of CAD associated gene SMAD3, a central transcription factor (TF) in the TGFβ pathway, investigating its role in smooth muscle biology. In vitro studies in human coronary artery smooth muscle cells (HCASMC) revealed that SMAD3 modulates cellular phenotype, promoting expression of differentiation marker genes while inhibiting proliferation. RNA sequencing and chromatin immunoprecipitation sequencing studies in HCASMC identified downstream genes that reside in pathways which mediate vascular development and atherosclerosis processes in this cell type. HCASMC phenotype, and gene expression patterns promoted by SMAD3 were noted to have opposing direction of effect compared to another CAD associated TF, TCF21. At sites of SMAD3 and TCF21 colocalization on DNA, SMAD3 binding was inversely correlated with TCF21 binding, due in part to TCF21 locally blocking chromatin accessibility at the SMAD3 binding site. Further, TCF21 was able to directly inhibit SMAD3 activation of gene expression in transfection reporter gene studies. In contrast to TCF21 which is protective toward CAD, SMAD3 expression in HCASMC was shown to be directly correlated with disease risk. We propose that the pro-differentiation action of SMAD3 inhibits dedifferentiation that is required for HCASMC to expand and stabilize disease plaque as they respond to vascular stresses, counteracting the protective dedifferentiating activity of TCF21 and promoting disease risk.

尽管通过全基因组关联研究（genome wide association studies）已将众多遗传基因座与冠状动脉疾病（coronary artery disease, CAD）建立关联，但目前仍需开展相关工作以识别这些基因座中的致病基因，并将其整合至核心信号通路之中。既往已有研究针对CAD相关基因SMAD3的疾病致病机制展开探索：该基因是转化生长因子β（transforming growth factor β, TGFβ）通路中的核心转录因子（transcription factor, TF），相关研究探讨了其在平滑肌生物学中的作用。针对人冠状动脉平滑肌细胞（human coronary artery smooth muscle cells, HCASMC）的体外实验显示，SMAD3可调控细胞表型，在促进分化标志物基因表达的同时抑制细胞增殖。对HCASMC开展的RNA测序（RNA sequencing）与染色质免疫沉淀测序（chromatin immunoprecipitation sequencing）研究，已鉴定出该细胞类型中参与介导血管发育与动脉粥样硬化过程的下游基因。研究发现，SMAD3所调控的HCASMC表型与基因表达模式，与另一CAD相关转录因子TCF21的作用方向截然相反。在DNA上SMAD3与TCF21共定位的区域，SMAD3的结合与TCF21的结合呈负相关，这在一定程度上是因为TCF21会在局部阻断SMAD3结合位点处的染色质可及性。此外，在转染报告基因（transfection reporter gene）实验中，TCF21可直接抑制SMAD3对基因表达的激活作用。与对CAD具有保护作用的TCF21不同，HCASMC中SMAD3的表达水平与疾病风险呈正相关。本研究提出，SMAD3的促分化作用可抑制HCASMC扩增及稳定病变斑块所需的去分化过程——该过程是细胞应对血管应激所必需的，从而抵消TCF21的保护性去分化活性，并最终提升疾病风险。