Department of Biochemistry and Molecular Biology Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA, USA.
BMC Cancer. 2019 Mar 22;19(1):258. doi: 10.1186/s12885-019-5454-2.
The oncoprotein MYC has the dual capacity to drive cell cycle progression or induce apoptosis, depending on the cellular context. BAG1 was previously identified as a transcriptional target of MYC that functions as a critical determinant of this cell fate decision. The BAG1 protein is expressed as multiple isoforms, each having an array of distinct biochemical functions; however, the specific effector function of BAG1 that directs MYC-dependent cell survival has not been defined.
In our studies the human osteosarcoma line U2OS expressing a conditional MYC-ER allele was used to induce oncogenic levels of MYC. We interrogated MYC-driven survival processes by modifying BAG1 protein expression. The function of the separate BAG1 isoforms was investigated by depleting cells of endogenous BAG1 and reintroducing the distinct isoforms. Flow cytometry and immunoblot assays were performed to analyze the effect of specific BAG1 isoforms on MYC-dependent apoptosis. These experiments were repeated to determine the role of the HSP70 chaperone complex in BAG1 survival processes. Finally, a proteomic approach was used to identify a set of specific pro-survival proteins controlled by the HSP70/BAG1 complex.
Loss of BAG1 resulted in robust MYC-induced apoptosis. Expression of the larger isoforms of BAG1, BAG1L and BAG1M, were insufficient to rescue survival in cells with oncogenic levels of MYC. Alternatively, reintroduction of BAG1S significantly reduced the level of apoptosis. Manipulation of the BAG1S interaction with HSP70 revealed that BAG1S provides its pro-survival function by serving as a cofactor for the HSP70 chaperone complex. Via a proteomic approach we identified and classified a set of pro-survival proteins controlled by this HSP70/BAG1 chaperone complex that contribute to the BAG1 anti-apoptotic phenotype.
The small isoform of BAG1, BAG1S, in cooperation with the HSP70 chaperone complex, selectively mediates cell survival in MYC overexpressing tumor cells. We identified a set of specific pro-survival clients controlled by the HSP70/BAG1S chaperone complex. These clients define new nodes that could be therapeutically targeted to disrupt the survival of tumor cells driven by MYC activation. With MYC overexpression occurring in most human cancers, this introduces new strategies for cancer treatment.
癌蛋白 MYC 具有双重能力，可以根据细胞环境推动细胞周期进程或诱导细胞凋亡。BAG1 先前被鉴定为 MYC 的转录靶标，是决定这种细胞命运决定的关键决定因素。BAG1 蛋白表达为多种异构体，每种异构体都具有一系列不同的生化功能；然而，指导 MYC 依赖性细胞存活的 BAG1 的特定效应功能尚未确定。
在我们的研究中，使用表达条件性 MYC-ER 等位基因的人骨肉瘤细胞系 U2OS 诱导致癌水平的 MYC。我们通过改变 BAG1 蛋白表达来研究 MYC 驱动的存活过程。通过耗尽细胞内源性 BAG1 并重新引入不同的异构体来研究单独的 BAG1 异构体的功能。通过流式细胞术和免疫印迹分析来分析特定 BAG1 异构体对 MYC 依赖性细胞凋亡的影响。这些实验被重复以确定 HSP70 伴侣复合物在 BAG1 存活过程中的作用。最后，采用蛋白质组学方法鉴定一组由 HSP70/BAG1 复合物控制的特定抗凋亡蛋白。
BAG1 的缺失导致强烈的 MYC 诱导的凋亡。表达 BAG1 的较大异构体 BAG1L 和 BAG1M 不足以挽救 MYC 致癌水平下细胞的存活。相反，BAG1S 的重新引入显著降低了细胞凋亡水平。操纵 BAG1S 与 HSP70 的相互作用表明，BAG1S 通过充当 HSP70 伴侣复合物的辅助因子来发挥其抗凋亡功能。通过蛋白质组学方法，我们鉴定并分类了一组由 HSP70/BAG1 伴侣复合物控制的特定抗凋亡蛋白，这些蛋白有助于 BAG1 的抗凋亡表型。
BAG1 的小异构体 BAG1S 与 HSP70 伴侣复合物合作，选择性地介导 MYC 过表达肿瘤细胞中的细胞存活。我们鉴定了一组由 HSP70/BAG1S 伴侣复合物控制的特定抗凋亡客户。这些客户定义了新的节点，可以通过治疗靶向来破坏由 MYC 激活驱动的肿瘤细胞的存活。由于大多数人类癌症中都存在 MYC 过表达，这为癌症治疗引入了新的策略。
