Flygare J, Armstrong R C, Wennborg A, Orsan S, Hellgren D
Department of Biosciences, CNT, Novum, Karolinska Institutet, Huddinge, Sweden.
FEBS Lett. 1998 May 8;427(2):247-51. doi: 10.1016/s0014-5793(98)00433-5.
The Rad51 gene of Saccharomyces cerevisiae is required for genetic recombination and recombinational repair of DNA strand breaks. In higher eukaryotes Rad51 is essential for embryonic development, and is involved in cell proliferation and DNA repair. Here we show that human Rad51 (HsRad51) is proteolytically cleaved during apoptosis in two T-lymphocyte cell lines, Jurkat and PFI-285. Apoptosis was induced by camptothecin or anti-Fas monoclonal antibody (anti-Fas mAb). HsRad51 was cleaved with similar kinetics as human poly(ADP-ribose) polymerase (HsPARP) after treatment with either agent. The time course of cleavage coincided with internucleosomal DNA fragmentation. The HsRad51 fragments observed in apoptotic cells were identical to those generated from in vitro translated (IVT) HsRad51 exposed to activated Jurkat S-100 extract in a cell-free system. In each case, cleavage of HsRad51 was abolished by acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO). However, cleavage of IVT HsRad51 could not be demonstrated using purified caspase-2, -3 or -6 to -10, and the identity of the responsible protease thus remains to be determined. In summary, we have shown that HsRad51 belongs to a group of repair proteins, including PARP and DNA-dependent protein kinase, which are specifically cleaved during the execution phase of apoptosis.
酿酒酵母的Rad51基因对于DNA链断裂的遗传重组和重组修复是必需的。在高等真核生物中，Rad51对于胚胎发育至关重要，并参与细胞增殖和DNA修复。在此我们表明，在两种T淋巴细胞系Jurkat和PFI - 285的凋亡过程中，人Rad51（HsRad51）会被蛋白水解切割。凋亡由喜树碱或抗Fas单克隆抗体（抗Fas mAb）诱导。用这两种试剂处理后，HsRad51的切割动力学与人聚（ADP - 核糖）聚合酶（HsPARP）相似。切割的时间进程与核小体间DNA片段化一致。在凋亡细胞中观察到的HsRad51片段与在无细胞系统中暴露于活化的Jurkat S - 100提取物的体外翻译（IVT）HsRad51产生的片段相同。在每种情况下，乙酰 - 天冬氨酸 - 谷氨酸 - 缬氨酸 - 天冬氨酸 - 醛（Ac - DEVD - CHO）都能消除HsRad51的切割。然而，使用纯化的半胱天冬酶 - 2、 - 3或 - 6至 - 10无法证明IVT HsRad51的切割，因此负责的蛋白酶的身份仍有待确定。总之，我们已经表明HsRad51属于一组修复蛋白，包括PARP和DNA依赖性蛋白激酶，它们在凋亡的执行阶段会被特异性切割。
