BCL-XL regulates the timing of mitotic apoptosis independently of BCL2 and MCL1 compensation

Mitotic catastrophe induced by prolonged mitotic arrest is a major Anticancer strategy. Although antiapoptotic BCL2-like proteins, including Bcl-xL, are known to regulate Apoptosis during mitotic arrest, adaptive changes in their expression can complicate loss-of-function studies. Our studies revealed compensatory alterations in the expression of BCL2 and MCL1 when Bcl-xL is either downregulated or overexpressed. To circumvent their reciprocal regulation, we utilized a degron-mediated system to acutely silence Bcl-xL just before mitosis. Our results show that in epithelial cell lines including HeLa and RPE1, Bcl-xL and BCL2 acted collaboratively to suppress Apoptosis during both unperturbed cell cycle and mitotic arrest. By tagging Bcl-xL and BCL2 with a common epitope, we estimated that Bcl-xL was less abundant than BCL2 in the cell. Nonetheless, Bcl-xL played a more prominent antiapoptotic function than BCL2 during interphase and mitotic arrest. Loss of Bcl-xL led to mitotic cell death primarily through a BAX-dependent process. Furthermore, silencing of Bcl-xL led to the stabilization of MCL1, which played a significant role in buffering Apoptosis during mitotic arrest. Nevertheless, even in a MCL1-deficient background, depletion of Bcl-xL accelerated mitotic Apoptosis. These findings underscore the pivotal involvement of Bcl-xL in controlling timely Apoptosis during mitotic arrest, despite adaptive changes in the expression of other BCL2-like proteins.