Knockdown of FAM3B triggers cell apoptosis through p53-dependent pathway
Knockdown of FAM3B triggers cell apoptosis through p53-dependent pathway
复制标题
FAM3B 的敲低通过 p53 依赖性途径触发细胞凋亡。
DOI：
10.1016/j.biocel.2012.12.003
复制
发表时间：
2013-03-01
影响因子：
4
通讯作者：
Le, Yingying
中科院分区：
文献类型：
--
作者：
Mou, Haiwei;Li, Zongmeng;Le, Yingying
FAM 3B，也称为PANDER，是2002年发现的一种类精氨酸蛋白。先前的研究表明，FAM 3B通过与肝脏和内分泌胰腺的相互作用来调节葡萄糖和脂质代谢。FAM 3B也在其他组织中表达，但其基本功能尚不清楚。本研究发现FAM 3B在小鼠结肠、肠、肝和肺组织以及多种类型的细胞系中表达，包括小鼠胰腺β细胞（Min 6）、小胶质细胞（N9）和肌细胞（C2 C12）;人结肠癌细胞（HCT 8、HCT 116、HT 29）、肝细胞（HL-7702）、肝细胞癌细胞（SMMC-7721）和肺癌细胞（A549）。通过RNA干扰抑制FAM 3B表达可诱导HCT 8、HCT 116、A549、N9、C2 C12和Min 6细胞的凋亡性细胞死亡，并降低HL-7702和小鼠原代肝细胞的细胞活力。进一步的研究表明，FAM 3B基因的敲除可增加膜结合Fas和Bax的蛋白水平，降低Bcl-2的表达，促进caspase-8、-3、-9和PARP的切割，并促进切割后PARP的核转位。这些结果表明，FAM 3B沉默激活外源性和内源性凋亡途径。机制研究表明，抗Fas中和抗体或沉默Fas相关死亡结构域对FAM 3B基因敲低诱导的细胞凋亡没有影响，而半胱天冬酶抑制剂可以显著逆转FAM 3B基因敲低诱导的细胞凋亡，表明Fas和死亡受体介导的外源性细胞凋亡途径不参与FAM 3B沉默诱导的细胞凋亡。进一步的研究表明，在FAM 3B敲低后，p53显著上调。沉默p53几乎可以完全逆转FAM 3B敲低诱导的Bax上调、Bcl-2下调、caspase-8、-9、-3切割和凋亡性细胞死亡，表明p53依赖性通路在FAM 3B沉默诱导的细胞凋亡中起关键作用。对HCT 116细胞的研究证实，FAM 3B表达的抑制通过p53依赖性途径诱导凋亡。此外，FAM 3B的敲低降低了Mdm 2的蛋白水平并促进了p53磷酸化。总之，我们的研究表明，沉默FAM 3B促进p53磷酸化，并通过降低Mdm 2表达诱导p53积累，这导致凋亡性细胞死亡。(c)2012爱思唯尔有限公司保留所有权利。
FAM3B, also named PANDER, is a cytokine-like protein identified in 2002. Previous studies showed that FAM3B regulates glucose and lipid metabolism through interaction with liver and endocrine pancreas. FAM3B is also expressed by other tissues but its basic function is unclear. In this study, we found that FAM3B was expressed in mouse colon, intestine, liver and lung tissues and multiple types of cell lines, including murine pancreatic beta-cell (Min6), microglia (N9) and muscle cell (C2C12); human colon cancer cells (HCT8, HCT116, HT29), hepatocyte (HL-7702), hepatocellular carcinoma cell (SMMC-7721) and lung carcinoma cell (A549). Inhibition of FAM3B expression by RNA interference induced apoptotic cell death of HCT8, HCT116, A549, N9, C2C12 and Min6 cells and decreased cell viability of HL-7702 and murine primary hepatocytes. Further studies with HCT8 cells showed that knockdown of FAM3B increased the protein levels of membrane-bound Fas and Bax, reduced the expression of Bcl-2, promoted the cleavage of caspases-8, -3, -9 and PARP, and the nuclear translocation of cleaved PARP. These results suggest that FAM3B silencing activates both extrinsic and intrinsic apoptotic pathways. Mechanistic studies showed that neutralizing antibody against Fas or silencing Fas-associated death domain had no effect on, while caspase inhibitors could significantly reverse FAM3B knockdown induced apoptosis, suggesting Fas and death receptor mediated extrinsic apoptotic pathway is not involved in FAM3B silencing induced apoptosis. Further studies showed that p53 was significantly upregulated after FAM3B knockdown. Silencing p53 could almost completely reverse FAM3B knockdown induced upregulation of Bax, downregulation of Bcl-2, cleavage of caspases-8, -9, -3, and apoptotic cell death, suggesting p53-dependent pathway plays critical roles in FAM3B silencing induced apoptosis. Studies with HCT116 cells confirmed that inhibition of FAM3B expression induced apoptosis through p53-dependent pathway. Furthermore, knockdown of FAM3B reduced the protein level of Mdm2 and promoted p53 phosphorylation. Taken together, our studies demonstrated that silencing FAM3B promoted p53 phosphorylation and induced p53 accumulation by decreasing Mdm2 expression, which resulted in apoptotic cell death. (c) 2012 Elsevier Ltd. All rights reserved.