Leonardi C L, Warren R H, Rubin R W
Biochim Biophys Acta. 1982 Apr 29;720(2):154-62. doi: 10.1016/0167-4889(82)90007-6.
We have utilized epithelial rat kidney cells and their Kirsten viral transformant (442) to examine the role of actin-binding proteins in cellular morphogenesis. Normal rat kidney cells are well spread while the transformed cells are more spherical, poorly adherent, and lack actin stress fibers (Rubin, R.W., Warren, R.H., Lukeman, D.S. and Clements, E. (1978) J. Cell Biol. 78, 28-35). By immunofluorescence, antitropomyosin prominently stains normal rat kidney cell stress fibers while only a weak, nonspecific fluorescence is observed in 442 cells. Using two-dimensional gel electrophoresis, tropomyosin can be detected in normal rat kidney cells homogenates. The tropomyosin subunits are enriched in Triton-extracted filamentous normal rat kidney cell models, and in extracts of normal rat kidney cell homogenate produced by using a rapid myosin affinity technique to isolate actin and actin-associated proteins. The identity of the tropomyosin subunits has been confirmed by electrophoretic mobility, lack of proline, and the peptide map generated by limited proteolysis. None of these techniques have detected tropomyosin in the corresponding 442 preparations. Our results suggest that the transformation of normal rat kidney cells has led to an overall reduction in tropomyosin content. This may be related to the inability of 442 cells to organize filamentous actin stress fibers.
我们利用大鼠肾上皮细胞及其柯斯顿病毒转化细胞（442）来研究肌动蛋白结合蛋白在细胞形态发生中的作用。正常大鼠肾细胞铺展良好，而转化细胞则更呈球形，贴壁性差，且缺乏肌动蛋白应力纤维（鲁宾，R.W.，沃伦，R.H.，卢克曼，D.S.和克莱门茨，E.（1978年）《细胞生物学杂志》78卷，28 - 35页）。通过免疫荧光法，抗原肌球蛋白显著地使正常大鼠肾细胞的应力纤维染色，而在442细胞中仅观察到微弱的非特异性荧光。使用二维凝胶电泳，可以在正常大鼠肾细胞匀浆中检测到原肌球蛋白。原肌球蛋白亚基在经Triton提取的丝状正常大鼠肾细胞模型中以及在通过使用快速肌球蛋白亲和技术分离肌动蛋白和肌动蛋白相关蛋白所产生的正常大鼠肾细胞匀浆提取物中富集。原肌球蛋白亚基的身份已通过电泳迁移率、脯氨酸的缺乏以及有限蛋白酶解产生的肽图得以确认。这些技术均未在相应的442制剂中检测到原肌球蛋白。我们的结果表明，正常大鼠肾细胞的转化导致了原肌球蛋白含量的总体降低。这可能与442细胞无法组织丝状肌动蛋白应力纤维有关。
