Abstract
Lysine acetyltransferases (KATs) are exquisitely fine-tuned to target specific lysine residues on many proteins, including histones, with aberrant acetylation at distinct lysines implicated in different pathologies. However, researchers face a lack of molecular tools to probe the importance of site-specific acetylation events in vivo. Because of this, there can be a disconnect between the predicted in silico or in vitro effects of a drug and the actual observable in vivo response. We have previously reported on how an in vitro biochemical analysis of the site-specific effects of the compound C646 in combination with the KAT p300 can accurately predict changes in histone acetylation induced by the same compound in cells. Here, we build on this effort by further analyzing a number of reported p300 modulators, while also extending the analysis to correlate the effects of these drugs to developmental and phenotypical changes, utilizing cellular and zebrafish model systems. While this study demonstrates the utility of biochemical models as a starting point for predicting in vivo activity of multi-site targeting KATs, it also highlights the need for the development of new enzyme inhibitors that are more specific to the regulation of KAT activity in vivo.
Highlights
Histone post-translational modifications play a major role in regulating access to DNA, and as such are important in the regulation of gene transcription and DNA repair
In order to quantify site-specific changes in p300 histone acetylation activity, we utilized selected reaction monitoring (SRM), a quantitative MS-based approach that enables the simultaneous measurement of acetylation of multiple lysine residues [7,20,23,24]
To characterize p300 histone acetylation activity, we began by measuring the catalytic efficiency of p300 at multiple sites of histone H3/H4; we have previously shown that determining how the site-specific catalytic efficiency of an acetyltransferase changes in response to external factors, such as drug treatment, can be utilized as a predictor of how that factor will alter the selectivity of a KAT
Summary
Histone post-translational modifications play a major role in regulating access to DNA, and as such are important in the regulation of gene transcription and DNA repair Due to this biological role, a significant amount of research focuses on creating a better understanding of the proteins that regulate histone modifications. One of the most studied KATs, p300, is a prolific acetyltransferase, with mutations in p300 resulting in several diseases including neurological disorders and cancer [1,2,3,4,5]. Because of this association, correcting aberrant levels of acetylation could prove instrumental in treating the diseases associated with them.
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