Profiling the role of non-enzymatic acetylation in cancer

FARE Award Winner

Authors

• RA Kulkarni
• TT Zengeya
• AM Roberts
• JL Meier

Abstract

Protein acetylation is a posttranslational modification critical for regulation of diverse cellular processes such as gene expression, metabolism and cell death. Altered cellular acetylation, found in many cancers, can modulate key tumor suppression or oncogenic signaling pathways as well as metabolic responses. While these findings implicate acetylation as a major driver of cell transformation and proliferation, the enzymes regulating many cellular acetylation events remain unknown. It has been hypothesized that the acetylation of many proteins may occur via spontaneous enzyme-independent reactivity with the electrophilic thioester of acetyl-CoA; however current methods to study this phenomenon in vivo are limited. Here we report the development of an unbiased chemical proteomic method to investigate the non-enzymatic reactivity of acetyl-CoA in cellular environments. We have developed a chemical probe, p-NAC, that specifically identifies proteins susceptible to non-enzymatic acetylation. This probe incorporates the intrinsically reactive thioester of acetyl-CoA, but lacks the adenine base and phosphopantetheine arm required for enzymatic acetylation. Following reaction with cancer cell proteomes, p-NAC can be ligated to a chemical reporter using “click chemistry” facilitating identification of thioester-reactive proteins. Our initial results show that p-NAC is cell permeable and non-toxic. Furthermore it reports exclusively on non-enzymatic acetylation events as it is not a substrate for acetyltransferases. Our current efforts are aimed at studying how non-enzymatic acetylation drive cell signaling in vivo and harnessing the intrinsic thioester reactivity to reestablish the cellular acetylation levels in cancers caused by hypoacetylation.

Scientific Focus Area: Chemical Biology

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