SM22α对血管平滑肌细胞肌动蛋白聚合和交联的调节  

THE MOLECULAR MECHANISMS OF SM22α IN CYTOSKELETON REMODELING OF VASCULAR SMOOTH MUSCLE CELLS

作　　者：温进坤[1] 史建红[1] 郑斌[1] 孟芳[1] 韩梅[1] 

机构地区：[1]河北医科大学基础医学研究所,河北省医学生物技术重点实验室,河北石家庄050017

出　　处：《中国应用生理学杂志》2008年第4期393-397,I0009,共6页Chinese Journal of Applied Physiology

基　　金：国家自然科学基金资助项目(30770787,30670845);河北省自然科学基金资助项目(C2006000814)

摘　　要：目的:探讨平滑肌22alpha(SM22α)调节血管平滑肌细胞(VSMC)骨架重构的分子机制。方法:血清饥饿法诱导VSMC由合成型转化成收缩型,转染pEGFP-SM22α表达质粒后观察SM22α在细胞中的分布及其与肌动蛋白纤丝(F-actin)的定位关系;应用反义技术封闭内源性SM22α表达,蛋白分步提取和Western blot分析检测敲减SM22α基因表达对肌动蛋白单体G-actin聚合的影响;F-actin体外交联实验观察SM22α对F-actin交联成束的影响。结果:SM22α在细胞中的分布与F-actin相一致;抑制内源性SM22α表达后,细胞中的SMα-actin主要以可溶性单体G-actin形式存在;F-actin体外交联实验结果表明,GST-SM22α蛋白纯品可促进F-actin交联形成粗大、束状的应力纤维,而敲减内源性SM22α的细胞裂解液促进F-actin交联的活性明显降低。结论:SM22α是参与VSMC细胞骨架重构的调节蛋白,不仅可促进G-actin聚合形成F-actin,而且还可加速F-actin交联成束,在VSMC骨架重构过程中起着十分重要的作用。Aim： To investigate the molecular mechanisms of smooth muscle 22 alpha（SM22α）whereby cytoskeleton remodeling of vascular smooth muscle cells （VSMCs） is regulated. Methods： Synthetic （dedifferentiated） VSMCs were converted to contractile （differentiated） VSMCs by serum deprivation.Cells were transfected with pEGFP-SM22α, and localization of SM22α and its relationship with F-actin were observed through fluorescence microscopy. Fractional extraction of proteins and Western blotting were used to detect polymerization of SM α-actin in antisense-pcD2-SM22α-transfected VSMCs. Furthermore, effect of SM22α on F-actin cross-linking was observed by F-actin polymerization experiment. Results： Fluorescence microscopy showed that SM22α co-localized with F-actin in contractile VSMCs. Western blotting of protein extracts from F-/G-actin fractions revealed that polymerization of SM α-actin was lower in antisense-pcD2-SM22α-transfected VSMCs, in which SM a-actin mostly existed as soluble G-actin. Moreover, F- actin polymerization in vitro also showed that GST-SM22α could promote cross-linking of F-actin to form thick and bundled stress fibres, while extracts from VSMCs transfected with antisense-pcD2-SM22α could not effectively induce the polymerization of F-actin. Conclusion： SM22α acts as a modulator to participate in VSMC cytoskeleton remodeling. It can not only induce polymerization of G- actin to F-actin,but also promote cross-linking of F-actin to bundled stress fibres, indicating its vital role in cytoskeleton remodeling.

关 键 词：平滑肌22α 血管平滑肌细胞 细胞骨架 肌动蛋白 

分 类 号：Q78[生物学—分子生物学]