Interaction between the BAG1S isoform and HSP70 mediates the stability of anti-apoptotic proteins and the survival of osteosarcoma cells expressing oncogenic MYC

Abstract

Background: 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. Methods: 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. Results: 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. Conclusions: 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.