Apoptosis repressor with caspase recruitment domain promotes cell proliferation and phenotypic modulation through 14-3-3ε/YAP signaling in vascular smooth muscle cells.,Journal of Molecular and Cellular Cardiology

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Apoptosis repressor with caspase recruitment domain promotes cell proliferation and phenotypic modulation through 14-3-3ε/YAP signaling in vascular smooth muscle cells.
Journal of Molecular and Cellular Cardiology ( IF 4.9 ) Pub Date : 2020-08-06 , DOI: 10.1016/j.yjmcc.2020.08.003
Mengxin Liu ₁ , Tao Yu ₂ , Mengyang Li ₃ , Xinyu Fang ₁ , Bo Hou ₄ , Gaoli Liu ₅ , Jianxun Wang ₃

Affiliation

Aims

In response to vascular injury, vascular smooth muscle cells (VSMC) may change from a contractile phenotype to a proliferative phenotype and consequently become conducive to neointima formation. Apoptosis repressor with caspase recruitment domain (ARC) was initially discovered as an endogenous apoptosis inhibitor, but whether ARC plays a role in VSMCs and whether it can participate in the regulation of atherosclerosis are unknown.

Methods and results

Protein and mRNA levels of ARC in tissues and cells were detected by western blot and quantitative real-time PCR. Immunofluorescence staining was used to detect the protein location, and immunohistochemistry was used to detect protein expression in tissues. VSMC proliferation was analysed using Cell Counting Kit-8 (CCK-8) and EdU assays, while migration was assessed by Transwell assay. Mechanistically, the direct binding between two proteins was verified by immunoprecipitation. We found that ARC expression was stimulated in VSMCs during cell proliferation. Our results also showed that ARC promoted cell proliferation and induced phenotypic modulation of VSMCs in vitro and vivo. Mechanistic studies demonstrated that ARC increased the nuclear localization of Yes associated protein (YAP) by binding to 14-3-3ε and that ARC played a role in promoting cell proliferation and phenotypic modulation. Additionally, the transcription factor p53 negatively regulated ARC expression at the transcriptional level during cell proliferation and phenotypic modulation.

Conclusions

Our findings define a novel role for ARC in the phenotypic transition of proliferating VSMCs, which may provide a new strategy for regulating neointimal formation.

中文翻译：

具有胱天蛋白酶募集结构域的凋亡抑制因子通过14-3-3ε/ YAP信号在血管平滑肌细胞中促进细胞增殖和表型调节。

目的

响应血管损伤，血管平滑肌细胞（VSMC）可能从收缩表型变为增生表型，因此有利于形成新内膜。最初发现具有胱天蛋白酶募集结构域（ARC）的凋亡抑制因子是一种内源性凋亡抑制剂，但是尚不清楚ARC是否在VSMC中起作用以及是否可以参与动脉粥样硬化的调节。

方法与结果

通过蛋白质印迹和定量实时PCR检测组织和细胞中ARC的蛋白质和mRNA水平。免疫荧光染色用于检测蛋白质的位置，免疫组织化学用于检测组织中的蛋白质表达。使用细胞计数试剂盒8（CCK-8）和EdU测定法分析VSMC增殖，同时通过Transwell测定法评估迁移。从机理上讲，两种蛋白质之间的直接结合通过免疫沉淀来验证。我们发现ARC表达在细胞增殖过程中在VSMC中被刺激。我们的研究结果还表明，ARC在体外和体内均可促进VSMC的细胞增殖并诱导表型调节。机理研究表明，ARC通过结合14-3-3ε来增加Yes关联蛋白（YAP）的核定位，并且ARC在促进细胞增殖和表型调节中起作用。此外，在细胞增殖和表型调节过程中，转录因子p53在转录水平上对ARC表达产生负调控。