Proteolytic activation of protein kinase C delta by an ICE-like protease in apoptotic cells
Proteolytic activation of protein kinase C delta by an ICE-like protease in apoptotic cells
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DOI：
10.1002/j.1460-2075.1995.tb00305.x
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发表时间：
1995-12-15
期刊：
影响因子：
11.4
通讯作者：
Kufe, D
中科院分区：
文献类型：
--
作者：
Emoto, Y;Manome, Y;Kufe, D
这些研究表明，用电离辐射（IR）处理人U-937细胞与细胞质髓鞘碱性蛋白（MBP）激酶的激活有关。通过凝胶过滤和凝胶内激酶实验对该激酶进行了表征，支持40 kDa蛋白的活化，底物和抑制剂的研究进一步支持诱导蛋白激酶C （PKC）样活性，纯化蛋白的n端氨基酸测序结果表明该激酶具有PKC δ的内部区域。免疫印迹分析证实，IR暴露后，对照细胞中完整的78 kDa PKC δ被蛋白水解裂解为40 kDa c端片段。研究发现，IR诱导的PKC δ蛋白水解激活和核内溶解DNA片段都被Bcl-2和Bcl-x(L)阻断，这支持了与生理性细胞死亡（PCD）的关联。此外，PKC δ的切割发生在天冬氨酸附近的一个位点（QDN），与白介素-1 β转换酶（ICE）的蛋白水解激活相似。特异性四肽ICE抑制剂（YVAD）阻断了ir处理细胞中PKC δ的蛋白水解激活和核小体间DNA的断裂。这些发现表明PCD与一种ice样蛋白酶对PKC δ的蛋白水解激活有关。
These studies demonstrate that treatment of human U-937 cells with ionizing radiation (IR) is associated with activation of a cytoplasmic myelin basic protein (MBP) kinase. Characterization of the kinase by gel filtration and in-gel kinase assays support activation of a 40 kDa protein, Substrate and inhibitor studies further support the induction of protein kinase C (PKC)-like activity, The results of N-terminal amino acid sequencing of the purified protein demonstrate identity of the kinase with an internal region of PKC delta. Immunoblot analysis was used to confirm proteolytic cleavage of intact 78 kDa PKC delta in control cells to the 40 kDa C-terminal fragment after IR exposure, The finding that both IR-induced proteolytic activation of PKC delta and endonucleolytic DNA fragmentation are blocked by Bcl-2 and Bcl-x(L) supports an association with physiological cell death (PCD). Moreover, cleavage of PKC delta occurs adjacent to aspartic acid at a site (QDN) similar to that involved in proteolytic activation of interleukin-1 beta converting enzyme (ICE), The specific tetrapeptide ICE inhibitor (YVAD) blocked both proteolytic activation of PKC delta and internucleosomal DNA fragmentation in IR-treated cells. These findings demonstrate that PCD is associated with proteolytic activation of PKC delta by an ICE-like protease.