Proteolysis by calpains: A possible contribution to degradation of p53
Proteolysis by calpains: A possible contribution to degradation of p53
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DOI：
10.1128/mcb.17.5.2806
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发表时间：
1997-05-01
影响因子：
5.3
通讯作者：
Piechaczyk, M
中科院分区：
文献类型：
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作者：
Pariat, M;Carillo, S;Piechaczyk, M
p53是一种在肿瘤细胞中频繁突变的短寿命转录因子。几个实验室的工作已经表明，泛素-蛋白酶体途径可以在很大程度上解释p53的破坏，至少在特定的实验条件下。我们在这里报告，在体外，野生型p53是一个敏感的底物的毫钙蛋白酶和微钙蛋白酶，这是丰富的和普遍存在的细胞质蛋白酶。降解依赖于p53蛋白的构象。具有改变的三级结构的p53突变体显示出对钙蛋白酶的广泛的易感性，其中一些在很大程度上抵抗降解，而另一些则更敏感。这一结果表明，这里测试的不同突变体采用略微不同的构象，钙蛋白酶对这些构象敏感，但不能通过使用单克隆抗体如PAb 1620和PAb 240来区分。通过使用生理抑制剂钙蛋白酶抑制素抑制钙蛋白酶导致表达野生型p53的细胞中p53稳态水平升高。相反，钙离子载体激活钙蛋白酶导致哺乳动物细胞中p53的减少，并且这种作用被细胞渗透性钙蛋白酶抑制剂阻断。用p53和钙蛋白酶抑制蛋白表达载体共转染p53-null细胞系导致p53依赖性转录活性增加。两者合计，这些数据支持的想法，钙蛋白酶也可能有助于调节野生型p53蛋白水平在体内。
p53 is a short-lived transcription factor that is frequently mutated in tumor cells. Work by several laboratories has already shown that the ubiquitin-proteasome pathway can largely account for p53 destruction, at least under specific experimental conditions. We report here that, in vitro, wild-type p53 is a sensitive substrate for milli- and microcalpain, which are abundant and ubiquitous cytoplasmic proteases. Degradation was dependent on p53 protein conformation. Mutants of p53 with altered tertiary structure displayed a wide range of susceptibility to calpains, some of them being largely resistant to degradation and others being more sensitive. This result suggests that the different mutants tested here adopt slightly different conformations to which calpains are sensitive but that cannot be discriminated by using monoclonal antibodies such as PAb1620 and PAb240. Inhibition of calpains by using the physiological inhibitor calpastatin leads to an elevation of p53 steady-state levels in cells expressing wild-type p53. Conversely, activation of calpains by calcium ionophore led to a reduction of p53 in mammalian cells, and the effect was blocked by cell-permeant calpain inhibitors. Cotransfection of p53-null cell lines with p53 and calpastatin expression vectors resulted in an increase in p53-dependent transcription activity. Taken together, these data support the idea that calpains may also contribute to the regulation of wild-type p53 protein levels in vivo.