González-Rodriguez Agueda, Alba Javier, Zimmerman Valeri, Kozma Sara C, Valverde Angela M
Instituto de Investigaciones Biomédicas Alberto Sols (CSIC/UAM), C/ Arturo Duperier 4, 28029 Madrid, Spain.
Hepatology. 2009 Jul;50(1):216-29. doi: 10.1002/hep.22915.
The mammalian target of rapamycin (mTOR)/S6K1 signaling pathway controls cell growth and proliferation. To assess the importance of S6K1 in the balance between death and survival in the liver, we have generated immortalized hepatocyte cell lines from wild-type and S6K1-deficient (S6K1(-/-)) mice. In S6K1(-/-) hepatocytes, caspase-8 and the pro-apoptotic protein Bid were constitutively down-regulated as compared with wild-type. Moreover, S6K1(-/-) hepatocytes failed to respond to the apoptotic trigger of death receptor activation. Neither caspase-8 activation nor FLIP(L) degradation in response to tumor necrosis factor alpha (TNF-alpha) or anti-Fas antibody (Jo2) was observed in cells lacking S6K1. Downstream events such as Bid cleavage, cytochrome C release, caspase-3 activation, DNA laddering, as well as the percentage of apoptotic cells were attenuated as compared with wild-type. In addition, the anti-apoptotic protein Bclx(L) was down-regulated in TNF-alpha-treated or Jo2-treated wild-type hepatocytes, but this response was abolished in S6K1(-/-)cells. In vivo, S6K1-deficient mice were protected against concanavalin A-induced apoptosis. The withdrawal of growth factors strongly induced apoptosis in wild-type, but not in S6K1(-/-) hepatocytes. S6K1 deficiency did not decrease Bclx(L)/Bim ratio on serum withdrawal, thereby protecting cells from cytochrome C release and DNA fragmentation. At the molecular level, the lack of S6K1-mediated negative feedback decreased insulin receptor substrate-1 (IRS-1) serine phosphorylation, resulting in activation of survival pathways mediated by phosphatidylinositol 3-kinase/Akt and extracellular signal-regulated kinase (ERK). However, S6K1(-/-) hepatocytes underwent apoptosis on serum withdrawal in combination with phosphatidylinositol 3-kinase (PI3K) or ERK inhibitors.
This finding might explain the mechanism of resistance to mTOR inhibitors in cancer treatments and strongly suggests that the inhibition of S6K1 could protect against acute liver failure and, in combination with inhibitors that abrogate the sustained activation of Akt and ERK, could improve the efficacy of hepatocarcinoma (HCC) treatment.
雷帕霉素哺乳动物靶点（mTOR）/S6K1信号通路控制细胞生长和增殖。为了评估S6K1在肝脏细胞死亡与存活平衡中的重要性，我们从野生型和S6K1基因缺陷（S6K1(-/-)）小鼠中建立了永生化肝细胞系。与野生型相比，在S6K1(-/-)肝细胞中，半胱天冬酶-8（caspase-8）和促凋亡蛋白Bid持续下调。此外，S6K1(-/-)肝细胞对死亡受体激活的凋亡触发无反应。在缺乏S6K1的细胞中，未观察到对肿瘤坏死因子α（TNF-α）或抗Fas抗体（Jo2）的反应中caspase-8激活或FLIP(L)降解。与野生型相比，下游事件如Bid裂解、细胞色素C释放、caspase-3激活、DNA梯状条带以及凋亡细胞百分比均减弱。此外，抗凋亡蛋白Bclx(L)在TNF-α处理或Jo2处理的野生型肝细胞中下调，但在S6K1(-/-)细胞中此反应被消除。在体内，S6K1基因缺陷小鼠对刀豆蛋白A诱导的凋亡具有抗性。生长因子撤除强烈诱导野生型肝细胞凋亡，但对S6K1(-/-)肝细胞无此作用。S6K1缺陷在血清撤除时并未降低Bclx(L)/Bim比值，从而保护细胞免受细胞色素C释放和DNA片段化影响。在分子水平，缺乏S6K1介导的负反馈降低了胰岛素受体底物-1（IRS-1）丝氨酸磷酸化，导致磷脂酰肌醇3激酶/蛋白激酶B（PI3K/Akt）和细胞外信号调节激酶（ERK）介导的存活通路激活。然而，S6K1(-/-)肝细胞在血清撤除并联合磷脂酰肌醇3激酶（PI3K）或ERK抑制剂时发生凋亡。
这一发现可能解释了癌症治疗中对mTOR抑制剂耐药的机制，并强烈提示抑制S6K1可预防急性肝衰竭，且与消除Akt和ERK持续激活的抑制剂联合使用，可提高肝癌（HCC）治疗效果。
