David H. Koch Institute for Integrative Cancer Research at MIT, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Genes Dev. 2013 May 1;27(9):1003-15. doi: 10.1101/gad.211326.112. Epub 2013 Apr 25.
The retinoblastoma protein gene RB-1 is mutated in one-third of human tumors. Its protein product, pRB (retinoblastoma protein), functions as a transcriptional coregulator in many fundamental cellular processes. Here, we report a nonnuclear role for pRB in apoptosis induction via pRB's direct participation in mitochondrial apoptosis. We uncovered this activity by finding that pRB potentiated TNFα-induced apoptosis even when translation was blocked. This proapoptotic function was highly BAX-dependent, suggesting a role in mitochondrial apoptosis, and accordingly, a fraction of endogenous pRB constitutively associated with mitochondria. Remarkably, we found that recombinant pRB was sufficient to trigger the BAX-dependent permeabilization of mitochondria or liposomes in vitro. Moreover, pRB interacted with BAX in vivo and could directly bind and conformationally activate BAX in vitro. Finally, by targeting pRB specifically to mitochondria, we generated a mutant that lacked pRB's classic nuclear roles. This mito-tagged pRB retained the ability to promote apoptosis in response to TNFα and also additional apoptotic stimuli. Most importantly, induced expression of mito-tagged pRB in Rb(-/-);p53(-/-) tumors was sufficient to block further tumor development. Together, these data establish a nontranscriptional role for pRB in direct activation of BAX and mitochondrial apoptosis in response to diverse stimuli, which is profoundly tumor-suppressive.
视网膜母细胞瘤蛋白基因 RB-1 在三分之一的人类肿瘤中发生突变。其蛋白产物 pRB（视网膜母细胞瘤蛋白）在许多基本的细胞过程中作为转录共调节剂发挥作用。在这里，我们通过发现 pRB 通过直接参与线粒体凋亡来增强 TNFα 诱导的凋亡，从而报告了 pRB 在凋亡诱导中的非核作用。我们发现这种促凋亡活性甚至在翻译被阻断时，pRB 也能增强 TNFα 诱导的凋亡。这种促凋亡功能高度依赖 BAX，表明其在线粒体凋亡中的作用，因此，内源性 pRB 的一部分与线粒体持续相关。值得注意的是，我们发现重组 pRB 足以在线粒体或脂质体体外引发 BAX 依赖性通透。此外，pRB 在体内与 BAX 相互作用，并且可以在体外直接结合并构象激活 BAX。最后，通过将 pRB 特异性靶向线粒体，我们生成了一种突变体，该突变体缺乏 pRB 的经典核作用。这种带有线粒体标签的 pRB 仍然能够响应 TNFα 以及其他凋亡刺激物来促进凋亡。最重要的是，在 Rb(-/-);p53(-/-)肿瘤中诱导表达带有线粒体标签的 pRB 足以阻止肿瘤的进一步发展。总之，这些数据确立了 pRB 在直接激活 BAX 和线粒体凋亡以响应各种刺激物方面的非转录作用，这对肿瘤具有强烈的抑制作用。
