Involvement of protein kinase C-δ in DNA damage-induced apoptosis
Involvement of protein kinase C-δ in DNA damage-induced apoptosis
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DOI：
10.1038/sj.cdd.4400885
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发表时间：
2001-09-01
影响因子：
12.4
通讯作者：
Johnson, CL
中科院分区：
文献类型：
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作者：
Basu, A;Woolard, MD;Johnson, CL
我们先前已经证明，蛋白激酶C(PKC)信号转导通路通过DNA损伤剂顺式二氨基氯铂(CDDP)调节细胞死亡。在本研究中，我们研究了PKC如何影响cDDP诱导的DNA损伤触发的一系列事件。CDDP引起caspase-8、-9、-3、-7的激活和PKC增量的断裂。新型PKC Delta的选择性抑制剂Rottlerin可阻断cDDP诱导的caspase激活、蛋白水解性激活和细胞死亡。相反，常规PKCα和βI的抑制剂GO 6976不能阻止cDDP诱导的caspase激活和cDDP细胞毒性。在HeLa细胞中，PKC Delta分布于胞浆和含有线粒体的重膜(HM)部分。虽然caspase-8主要是胞浆的，但在HM组分中可以检测到少量的caspase-9、-7和-3。CDDP引起线粒体细胞色素c释放的时间依赖性增加，胞浆和膜相关caspase的加工，以及PKC增量的蛋白水解性切割。Rottlerin可减弱cDDP对细胞色素c的晚期释放，但不影响其早期释放。然而，它抑制了胞浆和HM组分中caspase的激活和PKC Delta的蛋白水解性切割。Rotlerin与cDDP同时加入或随后加入时，抗细胞凋亡作用明显，而在去掉cDDP后加入则无明显抗凋亡作用。因此，PKC Delta抑制剂在cDDP诱导的细胞死亡途径的早期阶段起作用，该途径先于caspase激活。
We have previously shown that the protein kinase C (PKC) signal transduction pathway regulates cell death by the DNA damaging agent cis-diamminedichloroplatinum(II) (cDDP). In the present study we have investigated how PKC influences the sequence of events that are triggered by cDDP-induced DNA damage. cDDP caused activation of caspases-8,-9,-3,-7 and cleavage of PKC delta. Rottlerin, a selective inhibitor of novel PKC delta, blocked activation of caspases, proteolytic activation of PKC delta and cell death induced by cDDP. In contrast, Go 6976, an inhibitor of conventional PKC alpha and betaI, did not prevent cDDP-induced caspase activation and cDDP cytotoxicity. In HeLa cells, PKC delta was distributed both in the cytosol and heavy membrane (HM) fraction containing mitochondria. While caspase-8 was primarily cytosolic, a small amount of caspases-9, -7 and -3 could be detected in the HM fraction. cDDP caused a time-dependent increase in Cytochrome c release from the mitochondria and processing of both cytosolic and membrane-associated caspases, as well as proteolytic cleavage of PKC delta. Rottlerin attenuated late but not early release of Cytochrome c by cDDP. It, however, inhibited activation of caspases and proteolytic cleavage of PKC delta in both cytosolic and HM fractions. The antiapoptotic effect of rottlerin was evident when it was added together with or following cDDP addition but not when added after cDDP was removed from the medium. Thus, the PKC delta inhibitor acts at an early stage of the cDDP-induced cell death pathway that precedes caspase activation.