In vitro translation of rat liver and Novikoff hepatoma cytokeratin mRNAs.
In vitro translation of rat liver and Novikoff hepatoma cytokeratin mRNAs.
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大鼠肝脏和 Novikoff 肝癌细胞角蛋白 mRNA 的体外翻译。
DOI：
10.1007/bf00233805
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发表时间：
1986
影响因子：
4.3
通讯作者：
Hnilica,LS
中科院分区：
文献类型：
--
作者：
Krajewska,WM;Schmidt,WN;Hnilica,LS
从正常大鼠肝脏和Novikoff腹水肝癌细胞中分离出细胞质poly(A)+RNA，使用兔网织红细胞裂解物系统进行体外翻译，并使用Novikoff肝癌主要细胞角蛋白p39、p49（一组肝细胞角蛋白C、D和E）和p56特异性抗体进行免疫沉淀分析翻译产物。通过二维聚丙烯酰胺凝胶电泳进一步证实了沉淀抗原的身份。仅Novikoff肝癌poly(A)+RNA含有编码p39细胞角蛋白的可翻译mRNA，而p49和p56细胞角蛋白均从正常大鼠肝脏和Novikoff肝癌poly(A)+RNA翻译。使用 p39 特异性单克隆抗体进行免疫沉淀也回收了大量的 p56 和 49K 细胞角蛋白，这可能是由于这些蛋白质在体外合成后立即与 p39 形成寡聚体。在用抗p56单克隆抗体进行实验后观察到类似的结果，其中在免疫沉淀物中回收到不与该抗体反应的p39。总体而言，从 NAH 或肝脏 Poly(A)+RNA 体外合成的细胞角蛋白的二维凝胶荧光图与之前报道的分离抗原和细胞角蛋白图谱非常相似。这些结果表明，明显的翻译后加工不太可能导致肝脏中观察到的细胞角蛋白的多样性。
Cytoplasmic poly(A)+RNA was isolated from normal rat liver and Novikoff ascites hepatoma cells, translatedin vitrousing rabbit reticulocyte lysate system and the translational products were assayed by immunoprecipitation with antibodies specific for Novikoff hepatoma principal cytokeratins p39, p49 (a group of hepatic cytokeratins C, D, and E) and p56. The identity of the precipitated antigens was further confirmed by two-dimensional polyacrylamide gel electrophoresis. Only the Novikoff hepatoma poly(A)+RNA contained translatable mRNA coding for the p39 cytokeratin while the p49 and p56 cytokeratins were translated from both the normal rat liver and Novikoff hepatoma poly(A)+RNAs. Immunoprecipitations employing monoclonal antibody specific for p39 also recovered significant quantities of p56 and 49K cytokeratins, presumably due to oligomeric associations of these proteins with p39 immediately afterin vitrosynthesis. Similar results were observed after experiments with anti-p56 monoclonal antibody in which p39, not reactive with this antibody, was recovered in immunoprecipitates. Overall, the two-dimensional gel fluorograms of cytokeratins synthesized in vitro from NAH or liver poly(A)+RNA are quite similar to isolated antigenic and cytokeratin profiles reported previously. These results suggest that overt posttranslational processing is not likely responsible for the diversity of cytokeratins observed in the liver.