Overview
The eukaryotic genome is regulated by a variety of epigenetic mechanisms that establish and maintain proper gene expression profiles to control cell identity and fate. One of these vital mechanisms is accomplished by chromatin, which is the packaging medium for genomic DNA. The chromatin polymer consists of individual nucleosomes in which the DNA is wrapped around an octamer of the canonical histone proteins H2A, H2B, H3, and H4. The histone proteins are highly post-translationally modified, and these modifications (PTMs) act as dynamic signals to delineate specific chromatin states. Importantly, when histone PTMs and other epigenetic processes are dysregulated, this leads to aging and diseases including cancer and metabolic and developmental disorders.
Our lab’s goal is to understand how the deposition, removal, and recognition of these PTMs are regulated and what downstream effects these PTMs have on DNA-mediated processes. In particular, we want to understand how metabolism is linked to genomic regulation via histone PTMs. Thus, we seek to elucidate mechanisms by which the metabolic state of the cell (e.g., acetyl-CoA level) is reported to the genome via chromatin (e.g., histone acetylation) to lead to changes in DNA transcription or repair. To do so, my lab utilizes a range of techniques across organic chemistry, peptide/protein chemistry, biochemistry, and molecular and cell biology.
Metabolite Biosensors for Live Cell Imaging
Fluorescent biosensors are powerful tools for studying cell signaling dynamics, giving us a window into these processes as they are occurring in live cells. We are developing new genetically encoded biosensors for imaging metabolites, such as acyl-CoAs. With these sensors, we can track changes in subcellular metabolite levels in real time as cells respond to stimuli such as nutrient changes or inhibitor treatments. For example, we developed the first genetically encoded fluorescent biosensor for acetyl-CoA (link to preprint) and demonstrated its use in live cells. The sensor plasmids are available on Addgene. We are now working on developing new platforms for engineering metabolite biosensors that will improve on the throughput of traditional approaches to making these sensors.
Histone Acylations
The expansion of mass spectrometric characterization of histone PTMs has led to a recent surge in the identification of histone acylations beyond acetylation (e.g., succinylation, lactylation). While initial evidence suggests that at least some of these acylations perform distinct chromatin regulatory roles from acetylation and one another, these functions remain poorly defined. We use biochemical and cellular methods to delineate the mechanisms by which non-canonical acylations impact chromatin structure and function. Using protein semi-synthesis, we can produce site-specifically modified nucleosomes with these different acyl marks. We can then use these as physiologically relevant substrates to study eraser enzyme function and to profile reader domain preferences. We complement this biochemical work with cell culture models. For example, we work on defining regulatory feedback loops between the metabolic pathways that produce these acyl-CoAs and the effects on gene expression from the corresponding histone acylation.
Sirtuins
We have an ongoing interest in the NAD+-dependent deacylases and mono-ADP-ribosyltransferases called sirtuins. In particular, we focus on SIRT6 and SIRT7, which are primarily nuclear-localized sirtuins and are known to deacylate histones within nucleosomes. We are interested in the activity of these sirtuins to remove acyl groups other than acetyl, which may provide acyl-specific epigenetic regulatory mechanisms. Intriguingly, SIRT6 and other sirtuins can act not just as eraser enzymes but also as writers of mono-ADP-ribosylation (mARylation). SIRT6’s mARylation activity has been linked to oxidative stress and DNA damage response. Our goal is to determine substrates of SIRT6 mARylates and how this modification acts to facilitate the genomic stress response on a biochemical level.