Fujita N, Nagahashi A, Nagashima K, Rokudai S, Tsuruo T
Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.
Oncogene. 1998 Sep 10;17(10):1295-304. doi: 10.1038/sj.onc.1202065.
Interleukin-2 (IL-2)-dependent T cell clone CTLL-2 underwent apoptosis by deprivation of IL-2 from culture medium. The decrease in the anti-apoptotic Bcl-XL protein level was observed during apoptosis after IL-2 withdrawal. We found that Bcl-XL protein was cleaved to produce two 18 kDa fragments during CTLL-2 cell apoptosis. When the activation of caspases was suppressed by overexpressing human Bcl-2 protein or by the addition of caspase inhibitors, cleavage of Bcl-XL protein was suppressed in vivo. Bcl-XL protein cleavage by incubation with apoptosed CTLL-2 cell lysate was suppressed by the caspase-3/CPP32-specific tetrapeptide inhibitor in vitro. Therefore, caspase-3/CPP32-like proteases were activated and involved in the cleavage of Bcl-XL protein during CTLL-2 cell apoptosis. We found that Bcl-XL protein was cleaved by caspase-3/CPP32 at two sites in the loop domain (i.e., HLAD61/S and SSLD76/A). The transfection of the carboxy-terminal 18 kDa Bcl-XL fragment increased the sensitivity to apoptosis. These results indicate that caspase-3/CPP32-like proteases cleaved anti-apoptotic Bcl-XL protein and resulted in accelerated apoptotic cell death.
依赖白细胞介素-2（IL-2）的T细胞克隆CTLL-2在培养基中缺乏IL-2时会发生凋亡。在IL-2撤除后的凋亡过程中，观察到抗凋亡蛋白Bcl-XL水平下降。我们发现，在CTLL-2细胞凋亡过程中，Bcl-XL蛋白被切割产生两个18 kDa的片段。当通过过表达人Bcl-2蛋白或添加半胱天冬酶抑制剂来抑制半胱天冬酶的激活时，Bcl-XL蛋白的切割在体内受到抑制。在体外，通过与凋亡的CTLL-2细胞裂解物孵育进行的Bcl-XL蛋白切割被半胱天冬酶-3/CPP32特异性四肽抑制剂抑制。因此，在CTLL-2细胞凋亡过程中，半胱天冬酶-3/CPP32样蛋白酶被激活并参与Bcl-XL蛋白的切割。我们发现，Bcl-XL蛋白在环结构域的两个位点（即HLAD61/S和SSLD76/A）被半胱天冬酶-3/CPP32切割。羧基末端18 kDa的Bcl-XL片段转染增加了细胞对凋亡的敏感性。这些结果表明，半胱天冬酶-3/CPP32样蛋白酶切割抗凋亡的Bcl-XL蛋白，导致凋亡细胞死亡加速。
