Department of Medical Biochemistry, University of Amsterdam, 1105 AZ Amsterdam, The Netherlands.
J Biol Chem. 2011 Dec 30;286(52):44336-43. doi: 10.1074/jbc.M111.308999. Epub 2011 Nov 2.
The three members of the NR4A orphan nuclear receptor subfamily Nur77, Nurr1, and NOR-1, regulate a variety of biological functions including vascular disease and metabolism. In this study, we identified Four and a half LIM domains protein-2 (FHL2) as a novel interacting protein of NR4A nuclear receptors by yeast two-hybrid screen and co-immunoprecipitation studies. Each of the four LIM domains of FHL2 can bind Nur77, and both the amino-terminal domain and the DNA binding domain of Nur77 are involved in the interaction between FHL2 and Nur77. FHL2 represses Nur77 transcriptional activity in a dose-dependent manner, and short hairpin RNA-mediated knockdown of FHL2 results in increased Nur77 transcriptional activity. ChIP experiments on the enolase3 promoter revealed that FHL2 inhibits the association of Nur77 with DNA. FHL2 is highly expressed in human endothelial and smooth muscle cells, but not in monocytes or macrophages. To substantiate functional involvement of FHL2 in smooth muscle cell physiology, we demonstrated that FHL2 overexpression increases the growth of these cells, whereas FHL2 knockdown results in reduced DNA synthesis. Collectively, these studies suggest that association of FHL2 with Nur77 plays a pivotal role in vascular disease.
NR4A 孤儿核受体亚家族的三个成员 Nur77、Nurr1 和 NOR-1 调节多种生物学功能，包括血管疾病和代谢。在这项研究中，我们通过酵母双杂交筛选和共免疫沉淀研究鉴定了四个半 LIM 结构域蛋白 2 (FHL2) 是 NR4A 核受体的一种新的相互作用蛋白。FHL2 的四个 LIM 结构域中的每一个都可以与 Nur77 结合，并且 Nur77 的氨基末端结构域和 DNA 结合结构域都参与了 FHL2 和 Nur77 之间的相互作用。FHL2 以剂量依赖的方式抑制 Nur77 的转录活性，并且短发夹 RNA 介导的 FHL2 敲低导致 Nur77 转录活性增加。烯醇酶 3 启动子上的 ChIP 实验表明，FHL2 抑制 Nur77 与 DNA 的结合。FHL2 在人内皮和平滑肌细胞中高度表达，但不在单核细胞或巨噬细胞中表达。为了证实 FHL2 在平滑肌细胞生理学中的功能参与，我们证明了 FHL2 过表达增加了这些细胞的生长，而 FHL2 敲低导致 DNA 合成减少。总之，这些研究表明，FHL2 与 Nur77 的结合在血管疾病中起着关键作用。
