Inhibition of cell proliferation and cell cycle progression by specific inhibition of basal JNK activity - Evidence that mitotic Bcl-2 phosphorylation is JNK-independent
Inhibition of cell proliferation and cell cycle progression by specific inhibition of basal JNK activity - Evidence that mitotic Bcl-2 phosphorylation is JNK-independent
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DOI：
10.1074/jbc.m304935200
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发表时间：
2004-03-19
影响因子：
4.8
通讯作者：
Chambers, TC
中科院分区：
文献类型：
--
作者：
Du, LH;Lyle, CS;Chambers, TC
丝裂原活化蛋白激酶的c-Jun NH2末端激酶(JNK)亚群在应激反应中起重要作用，但越来越多的证据表明JNK也在细胞增殖和存活中发挥作用。我们研究了使用SP600125或特定的反义寡核苷酸抑制JNK对细胞增殖和细胞周期进程的影响。与ERK、p38、细胞周期蛋白依赖性蛋白激酶1(CDK1)和CDK2相比，SP600125在体外和体内对JNK具有选择性。SP600125对JNK活性和KB-3细胞增殖均有抑制作用，且呈相同剂量依赖关系，提示抑制细胞增殖是JNK抑制作用的直接结果。SP600125对细胞增殖的抑制作用与G(2)-M期和细胞凋亡率的增加有关，但与p53或p21的诱导作用无关。JNK2的反义寡核苷酸对细胞增殖的抑制作用非常显著，而JNK1的反义寡核苷酸对细胞增殖的抑制作用不明显。野生型小鼠成纤维细胞对增殖抑制和诱导凋亡的反应类似，而c-jun(-/-)成纤维细胞对SP600125的作用不敏感，提示c-jun的JNK信号是细胞增殖所必需的。在同步化的KB-3细胞中的研究表明，SP600125通过S和G(2)-M期延迟了通过时间。相应地，JNK活性在S晚期升高，在G(2)期晚期达到峰值。在同步有丝分裂过程中，随着c-jun、h1组蛋白和bcl2的磷酸化，细胞周期蛋白B水平升高。SP600125可延长有丝分裂进程，抑制c-jun的磷酸化，但不抑制h1和bcl2的磷酸化。然而，CDK抑制剂罗斯科维汀抑制了有丝分裂过程中的Bcl2磷酸化。这些结果表明，JNK，更具体地说，JNK2亚型，在细胞增殖和细胞周期进程中起着关键作用。此外，确凿的证据表明，除JNK外的其他激酶，最有可能是CDK1或CDK1调节的激酶，负责有丝分裂中的Bcl-2磷酸化。
The c-Jun NH2-terminal kinase (JNK) subgroup of mitogen-activated protein kinases has been implicated largely in stress responses, but an increasing body of evidence has suggested that JNK also plays a role in cell proliferation and survival. We examined the effect of JNK inhibition, using either SP600125 or specific antisense oligonucleotides, on cell proliferation and cell cycle progression. SP600125 was selective for JNK in vitro and in vivo versus other kinases tested including ERK, p38, cyclin-dependent protein kinase 1 (CDK1), and CDK2. SP600125 inhibited JNK activity and KB- 3 cell proliferation with the same dose dependence, suggesting that inhibition of proliferation was a direct consequence of JNK inhibition. Inhibition of proliferation by SP600125 was associated with an increase in the G(2)-M and apoptotic fractions of cells but was not associated with p53 or p21 induction. Antisense oligonucleotides to JNK2 but not JNK1 caused highly significant inhibition of cell proliferation. Wild-type mouse fibroblasts responded similarly with proliferation inhibition and apoptosis induction, whereas c-jun(-/-) fibroblasts were refractory to the effects of SP600125, suggesting that JNK signaling to c-Jun is required for cell proliferation. Studies in synchronized KB- 3 cells indicated that SP600125 delayed transit time through S and G(2)-M phases. Correspondingly, JNK activity increased in late S phase and peaked in late G(2) phase. During synchronous mitotic progression, cyclin B levels increased concomitant with phosphorylation of c-Jun, H1 histone, and Bcl-2. In the presence of SP600125, mitotic progression was prolonged, and c-Jun phosphorylation was inhibited, but neither H1 nor Bcl-2 phosphorylation was inhibited. However, the CDK inhibitor roscovitine inhibited mitotic Bcl-2 phosphorylation. These results indicate that JNK, and more specifically the JNK2 isoform, plays a key role in cell proliferation and cell cycle progression. In addition, conclusive evidence is presented that a kinase other than JNK, most likely CDK1 or a CDK1-regulated kinase, is responsible for mitotic Bcl-2 phosphorylation.