CELLULAR-LOCALIZATION AND CELL-CYCLE REGULATION BY A TEMPERATURE-SENSITIVE P53-PROTEIN
CELLULAR-LOCALIZATION AND CELL-CYCLE REGULATION BY A TEMPERATURE-SENSITIVE P53-PROTEIN
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DOI：
10.1101/gad.5.2.151
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发表时间：
1991-02-01
影响因子：
10.5
通讯作者：
LEVINE, AJ
中科院分区：
文献类型：
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作者：
MARTINEZ, J;GEORGOFF, I;LEVINE, AJ
原代大鼠胚胎成纤维细胞通过p53突变体（在135个氨基酸上丙氨酸转变为缬氨酸）和ras转化。这种p53val135突变体对温度敏感，通过结合单克隆抗体PAb246检测到构象变化，该抗体在32.5℃下识别野生型蛋白或大部分p53val135。在37℃时，突变型和野生型p53构象形式在细胞中共存，而在39.5℃时，细胞中的大部分p53val135呈突变型构象，不被PAb246抗体识别。在39.5摄氏度时，突变体p53定位于细胞的细胞质中。在32.5摄氏度时，p53蛋白进入细胞核并阻止这些细胞的生长。在37℃时，突变型和野生型p53混合存在，G1期间，野生型p53蛋白与hsc70和突变型p53蛋白在细胞质中形成复合体。当细胞进入细胞周期的s期时，这种野生型蛋白进入细胞核。在32.5摄氏度时，细胞停止复制并在G1/S边界停止。在32.5℃下加热48小时后，91%的细胞处于细胞周期的G1阶段。在进入下一个G1期之前，s期细胞似乎对p53负调控的生长免疫。这些数据强烈表明，转化细胞中的突变p53蛋白将野生型p53蛋白隔离在hsc70-p53复合体中，该复合体在细胞周期G1阶段存在于细胞质中，而核野生型p53通常在细胞周期中调节细胞生长和进展。通过这种方式，突变型p53蛋白可以以反显性方式发挥作用，克服细胞中野生型p53等位基因和蛋白对生长的调节。
Primary rat embryo fibroblasts were transformed by a p53 mutant (alanine to valine change at amino acid 135) plus ras. This p53val135 mutant is temperature sensitive for a conformational change detected by the binding of a monoclonal antibody, PAb246, which recognizes the wild-type protein or the great majority of p53val135 at 32.5-degrees-C. At 37-degrees-C, both mutant and wild-type p53 conformational forms co-exist in the cells, while at 39.5-degrees-C, the majority of the p53val135 in the cell is in a mutant conformation not recognized by PAb246 antibody. At 39.5-degrees-C, the mutant p53 is localized in the cytoplasm of the cell. At 32.5-degrees-C, the p53 protein enters the nucleus and stops the growth of these cells. At 37-degrees-C where there is a mixture of mutant and wild-type p53, the wild-type p53 protein is in a complex with hsc70 and mutant p53 protein in the cytoplasm of the cell during G1. This wild-type protein enters the nucleus as the cells enter the S-phase of the cell cycle. At 32.5-degrees-C, the cells stop replication and arrest at the G1/S border. After 48 hr at 32.5-degrees-C, 91% of the cells are in the G1 fraction of the cell cycle. The S-phase cells appear to be immune to the p53 negative regulation of growth until they enter the next G1 period. These data strongly suggest that mutant p53 proteins in transformed cells act to sequester the wild-type p53 protein in an hsc70-p53 complex, which resides in the cytoplasm during the stage of the cell cycle, G1, when nuclear wild-type p53 would normally act to regulate cell growth and progression through the cycle. In this way, mutant p53 proteins can act in a trans-dominant fashion to overcome growth regulation by the wild-type p53 allele and protein in a cell.