Ponchel F, Milner J
YCRC p53 Research Group, University of York, Department of Biology, UK.
Br J Cancer. 1998 May;77(10):1555-61. doi: 10.1038/bjc.1998.256.
p53 is activated in response to DNA damage and functions in the maintenance of genetic integrity. Loss of p53 function because of mutation of the p53 gene is associated with over half all human cancers. Certain human p53 mutants are conformationally flexible in vitro and are temperature sensitive, with partial or complete recovery of wild-type (wt) properties at 32 degrees C. We have now tested the functional capacities of selected p53 mutants in vivo, by transfection into established human cell lines. Unexpectedly, we found that wt p53 can be temperature sensitive for transactivation of a co-transfected target gene in vivo. Flexible mutants retained varying degrees of functional capacity in transfected cells, and the recipient cell line appeared to be a significant determinant of both wt and mutant p53 function; importantly, two p53 null cell lines commonly used to study p53 function (Saos-2 and Hep3B) differed markedly in this latter respect. We also show that the p53 mutant V272M, which exhibits sequence-specific DNA binding in vitro, is nonetheless defective for transactivation and is unable to induce apoptosis in vivo. The valine 272 residue may thus be crucial for properties (other than sequence-specific DNA binding) that are important for p53 function(s) in vivo.
p53因DNA损伤而被激活，并在维持基因完整性方面发挥作用。由于p53基因突变导致的p53功能丧失与超过半数的人类癌症相关。某些人类p53突变体在体外具有构象灵活性且对温度敏感，在32摄氏度时野生型（wt）特性会部分或完全恢复。我们现在通过转染已建立的人类细胞系，在体内测试了所选p53突变体的功能能力。出乎意料的是，我们发现在体内，wt p53对共转染靶基因的反式激活可能对温度敏感。灵活的突变体在转染细胞中保留了不同程度的功能能力，并且受体细胞系似乎是wt和突变型p53功能的重要决定因素；重要的是，常用于研究p53功能的两种p53基因缺失细胞系（Saos-2和Hep3B）在这方面存在显著差异。我们还表明，在体外表现出序列特异性DNA结合的p53突变体V272M，在体内反式激活方面存在缺陷，并且无法诱导细胞凋亡。因此，缬氨酸272残基对于体内p53功能重要的（除序列特异性DNA结合之外的）特性可能至关重要。
