ANALYSIS OF HUMAN P53 PROTEINS AND MESSENGER-RNA LEVELS IN NORMAL AND TRANSFORMED-CELLS
ANALYSIS OF HUMAN P53 PROTEINS AND MESSENGER-RNA LEVELS IN NORMAL AND TRANSFORMED-CELLS
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DOI：
10.1111/j.1432-1033.1986.tb09449.x
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发表时间：
1986-02-03
期刊：
影响因子：
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通讯作者：
CRAWFORD, L
中科院分区：
文献类型：
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作者：
MATLASHEWSKI, G;BANKS, L;CRAWFORD, L
在多种人转化细胞和正常人包皮成纤维细胞中检测p53 mRNA和蛋白表达。p53 mRNA的稳态和可翻译水平在正常和转化的人类细胞中是相同的。从正常和转化的人细胞中通过mRNA编程体外合成p53，发现这些细胞中p53的原代结构存在异质性。细胞脉冲标记和人反应性抗p53抗体免疫沉淀分析表明，体外合成的p53类型与体内由SV80和COLO 320细胞合成的p53类型相对应。HeLa和正常包皮成纤维细胞的类似脉冲标记分析未检测到p53。由于在本研究中有必要使用来自癌症患者的抗p53血清来进行大部分免疫沉淀分析，因此我们进一步表征这些血清，以确定它们是否与一个或多个表位反应。p53度量。在大肠杆菌中合成了-半乳糖苷酶融合蛋白，并用于分析癌症患者产生的抗p53抗体。我们证明了抗血清含有针对p53分子的n端和c端区域的表位的抗体。
p53 mRNA and proteins were examined in a variety of human transformed cells and in normal human foreskin fibroblast cells. Both the steady-state and translatable levels of p53 mRNA were the same in normal and transformed human cells. In vitro synthesized p53, programmed by mRNA from normal and transformed human cells, revealed that there was heterogeneity in the primary structure of p53 from these cells. Pulse labeling of cells and immunoprecipitation analysis with a panel of human reactive anti-p53 antibodies demonstrated that the types of p53 synthesized in vitro corresponded to the types made in vivo from SV80 and COLO 320 cells. No p53 was detectable by similar pulse-labeling analysis of HeLa and normal foreskin fibroblast cells. Since it was necessary to use anti-p53 sera from cancer patients to carry out much of the immunoprecipitation analysis in this study we therefore further characterised these sera to determine if they reacted with one or more than one epitope. p53-.beta.-galactosidase fusion proteins were synthesized in Escherichia coli and used to analyse the anti-p53 antibodies produced by cancer patients. We demonstrate that the antisera contain antibodies directed against epitopes in both the N-terminal and C-terminal regions of the p53 molecule.