Protein Interaction Between p53 and Δ113p53 Is Required for the Anti-Apoptotic Function of Δ113p53
Protein Interaction Between p53 and Δ113p53 Is Required for the Anti-Apoptotic Function of Δ113p53
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p53 和 Delta 113p53 之间的蛋白质相互作用是 Delta 113p53 抗凋亡功能所必需的
DOI：
10.1016/j.jgg.2014.01.001
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发表时间：
2014-02-01
影响因子：
5.9
通讯作者：
Chen, Jun
中科院分区：
文献类型：
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作者：
Ou, Zhao;Yin, Le;Chen, Jun
斑马鱼Delta 113 p53是一种N端截短的p53亚型，是一种拮抗p53介导的凋亡活性的p53靶基因。有趣的是，Delta 113 p53不以显性负性方式作用于p53，而是通过差异调节p53靶基因表达来干扰p53功能，以保护细胞免于凋亡。以往的研究表明，过量表达的Delta 113 p53和p53蛋白形成复合物。然而，目前尚不清楚内源性p53和Delta 113 p53蛋白是否也相互作用，以及Delta 113 p53是否需要这种相互作用来抑制全长p53的凋亡活性。在这项研究中，我们使用了两种可用的斑马鱼p53抗体来解决这些问题。其中一个Zfp 53-N只识别全长p53，而另一个Zfp 53-A7 C10则同时检测全长p53和Delta 113 p53。使用Zfp 53-N进行免疫沉淀和Zfp 53-A7 C10进行检测，我们证明了内源性Delta 113 p53和由DNA损伤药物诱导的全长p53在体内形成复合物。此外，在我们产生的六个Delta 113 p53突变体中，在寡聚化结构域中具有不同的点突变，其中两个突变体未能与p53相互作用，并且失去了调节p53靶基因表达和抑制p53诱导的细胞凋亡的能力。然而，那些可以与p53相互作用的Delta 113 p53突变体保留了拮抗p53凋亡活性的能力。因此，我们的数据表明，Delta 113 p53和p53之间的蛋白质-蛋白质相互作用是Delta 113 p53的抗凋亡功能所必需的。另外，两个不能与p53相互作用的Delta 113 p53突变体也可用于研究Delta 113 p53的其他功能的机制。
Zebrafish Delta 113p53, an N-terminal truncated p53 isoform, is a p53-target gene that antagonises p53-mediated apoptotic activity. Interestingly, Delta 113p53 does not act on p53 in a dominant-negative manner, but rather interferes with the p53 function by differentially modulating p53-target gene expression to protect cells from apoptosis. Previous studies showed that over-expressed Delta 113p53 and p53 proteins formed a complex. However, it is not known whether endogenous p53 and Delta 113p53 proteins also interact with each other, and if this interaction is required for Delta 113p53 to inhibit the apoptotic activity of full-length p53. In this study, we used two available zebrafish p53 antibodies to address these questions. One, Zfp53-N, only recognises full-length p53, whereas the other, Zfp53-A7C10, detects both full-length p53 and Delta 113p53. Using Zfp53-N for immunoprecipitation and Zfp53-A7C10 for detection, we demonstrated that endogenous Delta 113p53 and full-length p53 induced by a DNA-damaging drug formed a complex in vivo. Furthermore, of the six Delta 113p53 mutants we generated with different point mutations in the oligomerisation domain, two failed to interact with p53 and lost the ability to modulate p53-target gene expression and inhibit p53-induced cell apoptosis. However, those Delta 113p53 mutants that could interact with p53 retained the ability to antagonise the apoptotic activity of p53. Therefore, our data demonstrated that protein-protein interaction between Delta 113p53 and p53 is essential for the anti-apoptotic function of Delta 113p53. In addition, the two Delta 113p53 mutants that failed to interact with p53 are also useful for the study of the mechanisms of other functions of Delta 113p53.