Down-regulation of CIBZ, a novel substrate of caspase-3, induces apoptosis
Down-regulation of CIBZ, a novel substrate of caspase-3, induces apoptosis
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DOI：
10.1074/jbc.m802257200
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发表时间：
2008-05-23
影响因子：
4.8
通讯作者：
Kawaichi, Masashi
中科院分区：
文献类型：
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作者：
Oikawa, Yu;Matsuda, Eishou;Kawaichi, Masashi
我们之前鉴定并鉴定了一种含BTB结构域的小鼠蛋白CIBZ（在人类中是ZBTB38），它与CtBP相互作用并结合甲基化的CpGs。然而，其生理功能尚不清楚。据报道，CtBP参与p53非依赖性程序性细胞死亡，我们在此研究CIBZ是否与细胞凋亡有关。我们发现CIBZ在增殖的C2C12细胞中高表达，但在血清饥饿诱导凋亡后其表达水平下降。通过膜联蛋白V/碘化丙啶标记的增加、caspase-3的激活和poly（adp -核糖）聚合酶的裂解来确定，通过小干扰RNA敲低C2C12细胞中的CIBZ可诱导细胞凋亡。CIBZ抑制还激活了caspase-7和caspase-9，表明CIBZ相关的凋亡是通过线粒体途径发生的。值得注意的是，p53（-/-）小鼠胚胎成纤维细胞中CIBZ的敲低也激活了caspase-3和poly（adp -核糖）聚合酶的裂解，表明CIBZ相关的凋亡是由p53独立通路介导的；然而，由于CIBZ-和ctbp相关的细胞凋亡都是共同的和不同的目标，我们得出结论，不止一种途径参与其中。最后，通过诱变和体外caspase切割实验，我们发现CIBZ是caspase-3的新底物，并鉴定了两个caspase-3识别位点。综上所述，CIBZ参与了小鼠细胞凋亡的负向调控，发挥了重要的作用。
We previously identified and characterized a murine BTB domain-containing protein, CIBZ (ZBTB38 in human), that interacts with CtBP and binds to methylated CpGs. However, its physiological function remained unknown. As CtBP is reportedly involved in p53-independent programmed cell death, we examine here whether CIBZ is associated with apoptosis. We found that CIBZ was highly expressed in proliferating C2C12 cells but that its expression levels decreased upon induction of apoptosis by serum starvation. Knockdown of CIBZ by small interfering RNA in C2C12 cells induced apoptosis, as determined by an increase of annexin V/propidium iodide labeling, activation of caspase-3, and cleavage of poly( ADP-ribose) polymerase. CIBZ inhibition also activated caspase-7 and caspase-9, suggesting that CIBZ-associated apoptosis occurs through the mitochondrial pathway. Notably, knockdown of CIBZ in p53(-/-) mouse embryonic fibroblast cells also activated caspase-3 and cleavage of poly( ADP-ribose) polymerase, indicating that CIBZ-associated apoptosis is mediated by a p53-independent pathway; however, because both common and distinct targets are regulated by CIBZ- and CtBP-associated apoptosis, we conclude that more than one pathway is involved. Finally, using mutagenesis and an in vitro caspase cleavage assay, we show that CIBZ is a novel substrate of caspase-3 and identify two caspase-3 recognition sites. These findings indicate, collectively, that CIBZ plays an important role by participating in the negative regulation of apoptosis in murine cells.