花生四烯酸诱导肌球蛋白磷酸化被抑制的平滑肌细胞迁移的信号传导途径  

Signaling Pathways of Smooth Muscle Cell Migration when Phosphorylaion of Myosin Light Chain Was Inhibited

作　　者：吴晶辉[1] 叶婷婷[1] 王洪辉[1] 周晓雷[1] 米达[1] 吴莲英[1] 王长晔[1] 郭威[1] 张伟英[1] 张晓东[1] 叶丽虹[1] 

机构地区：[1]南开大学生命科学学院,天津300071

出　　处：《中国生物化学与分子生物学报》2006年第12期984-990,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金资助项目(No.30170203)~~

摘　　要：在应用肌球蛋白轻链激酶特异抑制剂ML-7抑制了肌球蛋白轻链磷酸化后,花生四烯酸(arachidonicacid,AA)仍可诱导兔血管平滑肌细胞(SM3)发生迁移.为了进一步阐明其信号传导途径,应用多种信号抑制剂,采用免疫印迹、Boyden小室和提取细胞膜蛋白等实验方法,对上述迁移作用的信号传导途径进行了深入的研究.结果显示,PTX(Gi蛋白抑制剂)、U73122(PLC抑制剂)、staurosporine(PKC抑制剂)、PD98059(ERK1/2抑制剂)和SB203580(p38抑制剂)分别可拮抗上述AA诱导的SM3细胞迁移作用,而SP600125(JNK抑制剂)的作用较弱.免疫印迹结果显示,AA可提高SM3细胞中PKC(ε)、ERK1/2、p38和JNK信号的磷酸化水平,呈时间依赖性,PTX或U73122可抑制上述作用;staurosporine可抑制由AA引起的ERK1/2和JNK的磷酸化水平增强,但对p38的磷酸化水平无影响.还发现AA可促进PLCβ2的细胞膜移位,PTX可抑制其作用.上述结果表明,当肌球蛋白轻链的磷酸化被抑制后,AA可通过Gi蛋白的活化促进PLCβ2向细胞膜移位,进而通过激活PKC(ε)、ERK1/2、p38和JNK等信号转导途径而诱导SM3细胞发生迁移.Arachidonic acid （AA）was able to induce migration of rabbit blood vessel smooth muscle cells （SM3） when phosphorylaion of myosin light chain （MLC20） was inhibited by ML-7, a specific inhibitor of myosin light chain kinase （MLCK）. To demonstrate the signaling pathways, we investigated the mechanism of above process by using the methods of application of protein inhibitors, Western blot analysis, Boyden＇ s chamber assay and experiment of extracted cell membrane. The results showed that the above migration process was markedly diminished by PTX （inhibitor of Gi protein）, U73122 （inhibitor of PLC）, Staurosporine （inhibitor of PKC ）, PD98059 （inhibitor of ERK1/2） and SB203580（inhibitor of p38）, and lightly by SP600125 （inhibitor of JNK）. It was found that phosphorylatiion levels of ERK1/2, p38, JNK and protein kinase Cε（PKCε） were enhanced by AA in a manner of time-course, however, the effect was remarkably attenuated by PTX or U73122. The enhancement of phosphorylation levels of ERK1/2 and JNK was depressed by staurosporine, but the phosphorylation of p38 was not affected by it. We also found that AA resulted in membrane translocation of PLCβ2 in the cells, which was abolished by PTX. Taken together, it suggests that the signaling pathways are related to promoting the membrane translocation of PLCβ2 by AA-activated Gi protein, furthermore resulting in migration of SM3 cells through enhancing phosphorylation levels of PKC （E）, ERK1/2, p38 and JNK, when phosphorylation of MLC20 was inhibited.

关 键 词：花生四烯酸 平滑肌细胞 肌球蛋白轻链非磷酸化 细胞迁移 信号传导途径 

分 类 号：R363[医药卫生—病理学] Q25[医药卫生—基础医学]