Abstract 521: Mechanisms of chemoresistance: linking lysine acetylation to cell death.

Abstract

Abstract Breast tumors often possess regions of hypoxia in which tumor cells must either adapt metabolically to survive or die. Some tumor cells adapt and, through a process that is poorly understood, become chemoresistant. We are interested in defining the mechanisms by which this chemoresistance develops, with focus on the role of lysine acetylation_an enigmatic protein modification thought to play a role in tuning metabolism to the available nutrient supply. We posit that changes in lysine acetylation reflect metabolic adaptation to hypoxia and modulate apoptotic-signaling pathways. The importance of protein lysine acetylation in tumor cell death is evidenced by numerous clinical cancer trials testing the tumor-killing potential of deacetylase inhibitors (e.g., Varinostat). However, the molecular mechanisms by which lysine acetylation modulates cell death are poorly understood. Using a proteomics approach to characterize acetylation dynamics under conditions of hypoxic stress in breast cancer cells, we found a striking correlation between changes in acetylation and hypoxia sensitivity. Specifically, proteome-wide increases in protein acetylation were only observed in hypoxia-sensitive cell lines, likely reflecting their attempt to adapt metabolically to hypoxia. Moreover, these experiments revealed potential networks of acetylated proteins involved in the hypoxia response, including metabolic enzymes, oncogenes (e.g., c-myc), nutrient-responsive transcription factors, and 14-3-3ζ. Our previously published data suggested that acetylation of 14-3-3ζ, a small pro-survival phospho-binding protein, led to its release from protein complexes and consequently sensitized cells to apoptosis (1). Consistent with this idea, we observed a 12-fold hypoxia-induced increase in 14-3-3ζ acetylation in the hypoxia-sensitive cells (with no detectable change in 14-3-3ζ acetylation in resistant cells). Moreover, this acetylation correlated with a shift of 14-3-3ζ from high to low molecular weight gel filtration fractions, indicating a dissociation of 14-3-3ζ from protein complexes. Proteomics efforts to characterize the 14-3-3ζ interactome in sensitive cells revealed that hypoxia triggers the release of 14-3-3ζ from proteins involved in glucose metabolism, and shifts its binding to proteins involved in autophagy and starvation metabolism, implicating 14-3-3ζ as a regulator of metabolism under these conditions. Importantly, depletion of 14-3-3ζ with siRNA (thus perturbing its network of interactions) potently sensitized all breast cancer cell lines, including the most highly resistant cells, to hypoxia. Collectively, our data suggest that lysine acetylation plays an integral part in dictating the survival of cells in hypoxia. Moreover, we propose that 14-3-3ζ, and the pathways that govern its acetylation, are promising therapeutic targets to sensitize breast cancer cells to death. Citation Format: Vajira Weerasekara, Jeff Mortenson, Lisa Heppler, Joshua Lyon Andersen. Mechanisms of chemoresistance: linking lysine acetylation to cell death. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 521. doi:10.1158/1538-7445.AM2013-521