Spindle poisons and cell fate: a tale of two pathways

Abstract

Spindle poisons, such as paclitaxel and vinblastine, exert their potent anti-neoplastic effects through activation of the spindle assembly checkpoint (SAC), thereby arresting cells in mitosis. Unfortunately, only certain cancers are susceptible to these drugs, and many patients fail to respond to treatment. We review the pathways that are triggered by spindle poisons and highlight recent studies that describe the great variability of tumor cells in responding to these drugs. We also describe the recent identification of an apoptotic pathway that is activated by mitotic arrest in response to spindle poisons. Emerging from these studies is not only a greater understanding of how these classic antimitotic agents bring about cell death, but also a wealth of potential new targets of anticancer therapeutics.

Figure 1. The spindle assembly checkpoint (SAC) controls the metaphase to anaphase transition

By inhibiting the degradation of cyclin B by the anaphase promoting complex/cyclosome (APC/C), the SAC arrests the cell in metaphase until all kinetochores are attached to spindle microtubules. A. In the absence of microtubule attachments, unattached kinetochores recruit checkpoint proteins and activate them, allowing Mad2, Bub3, and BubR1 to inhibit Cdc20 activation of APC/C, resulting in high amounts cyclin B and metaphase arrest. B. When all kinetochores have proper microtubule attachments, checkpoint proteins are no longer recruited to kinetochores and Cdc20 activates APC/C, which drives cyclin B degradation. Depletion of cyclin B causes anaphase onset.

Figure 2. Cells display a surprising amount of within-group variation in response to treatment with the spindle poison nocodazole

Population level analyses reveal the percentage of HeLa cells that survive (blue) or apoptose (white) following treatment with nocodazole. Analysis of individual cells reveals the large amount of variation in how the cells die (smaller pie). Only a small minority of HeLa cells treated with nocodazole die during the first mitotic arrest, which is the presumed mode by which spindle toxins bring about cell death. Cells that survive represent both survival in interphase after mitosis (8%) and survival after replicating their genomes but failing to complete mitosis, i.e. endocycling (55%).

Figure 3. Mcl concentrations control apoptotic timing during mitotic arrest

During prolonged mitosis, Mcl1 prevents the induction of apoptosis by inhibiting the binding of Bak and Bax to the mitochondrial membrane. However, Mcl1 levels fall over time. The protein kinases p38, Jun N-terminal kinase (JNK), and casein kinase II (CKII) phosphorylate Mcl1, driving its interaction with FBW7, the substrate binding component of a ubiquitin ligase complex. Ubiquitinated Mcl1 is then degraded by the proteasome. The deubiquitinase USP9X and protein phosphatase PP2A can promote Mcl1 stability by removing ubiquitin side-chains and dephosphorylating Mcl1, respectively. When Mcl1 concentrations fall low enough, Bak and Bax form pores in the mitochondrial membrane, resulting in the release of cytochrome c and terminal caspase activation and in apoptosis.

Figure 4. The spindle assembly checkpoint (SAC) and the Mcl1 “apoptotic timer” compete for the fate of cells during a mitotic arrest

A. SAC activation leads to mitotic arrest and triggers the onset of Mcl1 degradation. Mcl1 is acted on by a set of pro-apoptotic proteins that drive its degradation and a set of pro-survival proteins that enhance its stability. During prolonged mitotic arrest, the pro-apoptotic pathway is favored. Simultaneously, incomplete inhibition of the APC/C results in slow cyclin B degradation, a phenomenon known as slippage. B. Cells treated with spindle poisons can exit mitosis if cyclin B concentrations fall below the threshold (dotted line) before the concentration of Mcl1 does. C. If Mcl1 levels fall below threshold (dotted line) before cyclin B, the apoptotic pathway is triggered while the cell is in mitosis.