Regulating Mitotic Fidelity and Susceptibility to Cell Death: Non-Canonical Functions of Two Kinases

Abstract

<p>In this dissertation, I will present two studies on the non-canonical role of the how two kinases participate in the oncogenesis via non-canonical mechanisms. In the first study, we investigate the role of CoA synthase (COASY) in the regulation of the protein acetylation during mitosis. In the second study, we will investigate the dysregulation of the RIPK3 in the recurrent tumor cells render them uniquely susceptible to death signaling triggered by cystine deprivation. </p><p>The temporal activation of kinases and timely ubiquitin-mediated degradation is central to faithful mitosis. However, whether acetylation of mitotic proteins are involved in regulation mechanism of mitosis is relatively rarely studied. Here we present evidence that acetylation controlled by COASY and acetyltransferase CBP constitutes a novel mechanism that ensures faithful mitosis. We found that COASY knockdown triggers prolonged mitosis and multinucleation. Acetylome analysis reveals that COASY inactivation leads to hyper-acetylation of a subset of proteins associated with mitosis, including CBP and an Aurora A kinase activator, TPX2. We found that TPX2 is hyper-acetylated under COASY knockdown, which rendered TPX2 accumulation by resistant to ubiquitination. Further experiments confirmed that the direct interaction of COASY and CBP regulates CBP-mediated TPX2 acetylation. We therefore propose a regulatory mechanism that a transient CBP-mediated TPX2 acetylation is associated with TPX2 accumulation and Aurora A activation during early mitosis. The recruitment of COASY inhibits CBP-mediated TPX2 acetylation, promoting TPX2 degradation for mitotic exit. Remarkably, pharmacological and genetic inactivation of CBP effectively rescued the mitotic defects caused by COASY knockdown. We also found that PPAT domain on COASY is in responsible for the inhibitory effect of COASY on CBP. Together, our findings uncover a novel mitotic regulation wherein COASY and CBP coordinate an acetylation network to enforce productive mitosis. </p><p>Tumor recurrence results in most of the mortality in breast cancer. Recurrent tumor cells are usually incurable and unresponsive to most of the treatments. We have previous reported that triple negative breast cancer cells are more sensitive to Cystine deprivation though epithelial-mesenchymal transition. Here we report that recurrent tumor cells are more sensitive to Cystine deprivation when comparing to primary tumor cells. RNA expression profiling identifies a much higher expression of RIPK3 RNA in recurrent cells. RIPK3 silencing in recurrent cells can rescue the programmed necrosis under Cystine deprivation. We further found that RIPK3 is crucial for the rapid cell proliferation in recurrent cells. Thus, the collateral vulnerability can be a potential therapeutic target for recurrent tumor specific treatment. We are still actively investigating the molecular mechanism for the recurrent cells taking advantage of high RIPK3 expression for rapid cell growth.</p>