Schmid G, Wang Z Q, Wesierska-Gadek J
Institute of Tumorbiology-Cancer Research, University of Vienna, Austria.
Biochem Biophys Res Commun. 1999 Feb 16;255(2):399-405. doi: 10.1006/bbrc.1999.0191.
We have investigated the effect of PARP gene inactivation on the expression of wild-type (wt) p53 protein. Using immortalized fibroblasts from control and PARP knock-out mice we have found by immunoblotting with the PAb421 antibody a profound decrease of the p53 expression to a barely detectable level in PARP knock-out cells. Surprisingly, longer exposure of immunoblots revealed an immunoreactive band at about 75 kD which was stronger in PARP-deficient cells than in wt cells and was not affected upon doxorubicin treatment. The size of the PAb421 immunoreactive protein and the lack of its inducibility in response to DNA damage resembled those of p73, the first described p53 homologue. Therefore, we examined the reactivity of anti-p53 antibodies with in vitro translated p73 protein. Interestingly, p73 was efficiently immunoprecipitated with distinct antibodies recognizing the carboxy-terminus of p53. In Northern blots we observed p73 signals of comparable intensity in controls and PARP-deficient cells. We conclude that elevated expression of p73 may compensate the reduced level of p53 in PARP-deficient cells.
我们研究了聚（ADP - 核糖）聚合酶（PARP）基因失活对野生型（wt）p53蛋白表达的影响。利用来自对照小鼠和PARP基因敲除小鼠的永生化成纤维细胞，我们通过用PAb421抗体进行免疫印迹分析发现，在PARP基因敲除细胞中，p53表达显著降低至几乎检测不到的水平。令人惊讶的是，免疫印迹的长时间曝光显示在约75kD处有一条免疫反应带，该条带在PARP缺陷细胞中比在野生型细胞中更强，并且在阿霉素处理后不受影响。PAb421免疫反应蛋白的大小及其对DNA损伤缺乏诱导性与p73相似，p73是最早描述的p53同源物。因此，我们检测了抗p53抗体与体外翻译的p73蛋白的反应性。有趣的是，p73能被识别p53羧基末端的不同抗体有效免疫沉淀。在Northern印迹中，我们在对照细胞和PARP缺陷细胞中观察到强度相当的p73信号。我们得出结论，p73表达的升高可能补偿了PARP缺陷细胞中p53水平的降低。
