血管平滑肌细胞（VSMC）表型转变是动脉粥样硬化及支架安置后再狭窄病理发生中的关键步骤。如何维持其收缩表型是防治粥样硬化的新思路。蛋白激酶G（PKG）及ROCK激酶是增加肌纤维合成从而阻止其表型转变的重要通路，二者下游有肌纤维合成的关键分子FHOD1。我们前期研究解明PKG能磷酸化FHOD1-1131S及FHOD1氨基端能特异结合Rac1，但二者的生理意义不明。在此基础上日本人报告了边上新位点1137S及1141T，三点都被ROCK磷酸化后能激活FHOD1。我们预实验发现FHOD1羧基端还有一个Rac1新结合域；突变体放射自显影揭示PKG只磷酸化1131S，ROCK2只磷酸化另两位点；随着磷酸化增多羧基端结合Rac1能力提高；大鼠颈动脉新生内膜中FHOD1表达下调；接下来要在细胞内证明Rac1新结合域；Rac1如何调节两激酶及其对VSMC表型转变的作用，从而提供维持收缩表型的新分子靶点。

Phenotype change of vascular smooth muscle cells (VSMC) from contractile to synthetic is a key step in the pathogenesis of atherosclerosis and post-stent re-stenosis of the coronary artery lumen due to the neointima formation. Restraining VSMC within the contractile phenotype as fully as possible is a novel approach for future prevention and treatment of atherosclerosis. Two famous signaling pathways, nitric oxide/ANP-sGC-cGMP-PKG, and RhoA-ROCK-actin-myocardin/MRTF-serum response factor, have been reported to be deeply involved in the process of keeping VSMC within the contractile phenotype by regulating actin stress fiber (SF) formation as well as a series of VSMC-specific protein expression, like SM-MHC, SM-alpha-actin, calponin, smoothelin, and SM22-alpha. In the downstream of both pathways, a key regulator of SF formation is formin homology domain-containing protein 1 (FHOD1), a member of formin in human. ..We have previously reported that FHOD1 is a substrate of PKG in VSMC. PKG phosphorylates FHOD1 1131S within the C-terminal auto-regulatory domain (DAD) and FHOD1 binds specially to Rac1, but not RhoA and Cdc42, in a GTP-independent way (Wang-YP, et al. J Biol Chem. 2004; 279:24420-6). However, the physiological significance of these two processes is still unclear. Four years later, on the basis of our report, a Japanese group reported that another two adjacent residues, 1137S and 1141T in the DAD, can be phosphorylated by ROCK. Only getting all three residues phosphorylated could almost fully activate FHOD1 (Takeya-R, et al. EMBO J. 2008; 27:618–28)...Our pilot studies include: 1) we discovered a novel Rac1 binding site in FHOD1 C-terminus, which is also GDP/GTP-independent; 2) P32 autoradiography phosphorylation assay of a series of FHOD1 phosphoresistent or phosphomimetic mutants by pure PKG, ROCK1, ROCK2 demonstrated that PKG only phosphorylates 1131S and that ROCK2, but not ROCK1, phosphorylates 1137S and 1141T; 3) As FHOD1 C-terminus gets more sites phosphorylated, it could recruit more Rac1; 4) FHOD1 is down-regulated in the neointima formed in the injured rat carotid artery. ..Next, we plan to: 1) ascertain both in vitro and in vivo that the FHOD1 C-terminus binding is specific to Rac1; 2) test whether Rac1-FHOD1 C-terminus interaction could regulate PKG/ROCK2 phosphorylation as well as FHOD1 activation; and 3) study whether Rac1-FHOD1 C-terminus interaction could regulate VSMC’s phenotype. Our study aims to investigate whether Rac1-FHOD1 C-terminus interaction is a new regulator of FHOD1 activation and a drug-discovery target for effectively restraining VSMC in the contractile phenotype.