Biology subjects

Haws, S. A.

Publications and source records attributed to Haws, S. A..

4 recordsLinked to original sources

bioRxiv2026

dCas9 metabolic enzyme fusions modulate global and locus specific gene expression

Central metabolites function as essential co-substrates for chromatin-modifying enzymes, directly linking cellular metabolism to chromatin regulation. Accordingly, whole-cell fluctuations in co-substrate availabilities have been shown to promote diverse phenotypes through chromatin-dependent mechanisms. There is emerging evidence that metabolic enzymes producing co-substrates for chromatin modifying enzymes can exist in the nucleus, suggesting that nucleus-specific metabolite availability regulates chromatin state. Here, we developed CRISPRm (CRISPR metabolite) to assess how nucleus-specific metabolic perturbations influence chromatin function. Five dCas9-metabolic enzyme fusions (i.e., dCas9-ACSS2, -NMNAT1, -MAT2A, -GDH, and -AHCY) were used to modulate nuclear levels of essential co-substrates involved in histone (de)acetylation and (de)methylation reactions. Transient expression of all dCas9 fusions in HEK293T cells induced distinct global changes in gene expression patterns, with dCas9-ACSS2 (acetyl-CoA producing) and NMNAT1 (NAD+ producing) eliciting large opposing changes in gene expression, suggesting transcriptional responses to nuclear acetyl-CoA and NAD+ production may be directly facilitated by acetylation or deacetylation reactions, respectively. Targeting dCas9-ACSS2 and -NMNAT1 to promoters of select candidate genes revealed enhanced transcriptional modulation. dCas9-ACSS2 upregulated, and dCas9-NMNAT1 downregulated genes showed basal enrichment of H3K9ac, H3K18ac, H3K27ac, H3K4me3, and p300, suggesting these genomic loci reside within epigenetic environments susceptible to fluctuations in acetyl-CoA and NAD+ availability. Of significant genes altered, dCas9-MAT2A (SAM producing) increased expression of 72% whereas dCAS9-GDH (alpha-ketoglutarate producing) decreased expression of 79%. Surprisingly, dCAS9-AHCY (SAH hydrolysis) led to down-regulation of shared genes up-regulated by dCas9-MAT2A. The observations amongst the methylation-specific enzymes revealed unexpected and unique gene-regulatory sensitivities to SAM, SAH and alpha-ketoglutarate. Together, these results demonstrate the utility of CRISPRm in studying nuclear metabolic regulation of transcription and provide strong evidence that perturbations in nuclear co-substrates do not lead to a large mass- action changes in chromatin acetylation/methylation but rather to modulation of select chromatin-modifying enzymes with targeted transcription responses. HighlightsO_LICRISPRm is a novel, modular dCas9-effector platform that enables interrogation of the metabolism-epigenome axis C_LIO_LIdCas9-ACSS2, -NMNAT1, -MAT2A, -AHCY, and -GDH induce distinct transcriptional programs. C_LIO_LITargeting CRISPRm to promoters enhances transcriptional responses. C_LIO_LIdCas9-ACSS2 and -NMNAT1 sensitive gene promoters exhibit unique enrichment of chromatin features, including H3K9ac, H3K18ac, H3K27ac, H3K4me3 and p300. C_LI

biochemistry↗

bioRxiv2026

Methionine, Not S-adenosylmethionine, Acts as a Primary Metabolic Stress Signal for Chromatin Remodeling

Epigenetic regulation is tightly linked to cellular metabolism through chromatin-modifying enzymes that depend on central metabolites as co-substrates. Methionine is an essential amino acid that is directly converted by methionine adenosyltransferase 2A (MAT2A) into S-adenosylmethionine (SAM), the universal methyl donor required for histone and DNA methylation. Although methionine restriction/depletion can alter the chromatin methylation landscape and improve physiological outcomes in diverse biological systems, it remains unclear whether these effects arise from loss of methionine itself or from secondary depletion of SAM. Here, we show that methionine depletion induces nuclear accumulation of MAT2A together with redistribution of H3K9 methylation, derepression of transposable elements, activation of stress-response pathways, and broad transcriptional reprogramming. Surprisingly, pharmacologic inhibition reduced intracellular SAM to levels comparable to methionine depletion but failed to reproduce these major epigenetic or transcriptional responses. Furthermore, depletion of the SAM-sensor SAMTOR and inhibition of KDM4 histone demethylases did not prevent methionine-dependent chromatin remodeling, indicating that canonical SAM-sensing pathways are not required for this adaptation. Instead, methionine depletion uniquely induced innate immune and integrated stress-response programs consistent with a viral mimicry-like state. These findings demonstrate that methionine availability, rather than SAM abundance, functions as a primary metabolic signal regulating epigenetic adaptation to nutrient stress. Our data support a model in which methionine is sensed independently of SAM abundance and acts upstream of stress signaling pathways that secondarily remodel chromatin.

biochemistry↗

bioRxiv2022

Intrinsic catalytic properties of histone H3 lysine-9 methyltransferases preserve monomethylation levels under low S-adenosylmethionine

S-adenosylmethionine (SAM) is the methyl donor for site-specific methylation reactions on histone proteins, imparting key epigenetic information. During SAM-depleted conditions that can arise from dietary methionine restriction, lysine di- and tri-methylation are reduced while sites such as Histone-3 lysine-9 (H3K9) are actively maintained, allowing cells to restore higher-state methylation upon metabolic recovery. Here, we investigated if the intrinsic catalytic properties of H3K9 histone methyltransferases (HMTs) contribute to this epigenetic persistence. We employed systematic kinetic analyses and substrate binding assays using four recombinant H3K9 HMTs (i.e., EHMT1, EHMT2, SUV39H1, and SUV39H2). At both high and low (sub-saturating) [SAM], all HMTs displayed the highest catalytic efficiency (kcat/KM) for monomethylation compared to di- and trimethylation on H3 peptide substrates. The favored monomethylation reaction was also reflected in kcat values, apart from SUV39H2 which displayed a similar kcat regardless of substrate methylation state. Using differentially-methylated nucleosomes as substrates, kinetic analyses of EHMT1 and EHMT2 revealed similar catalytic preferences. Orthogonal binding assays revealed only small differences in substrate affinity across methylation states, suggesting that catalytic steps dictate the monomethylation preferences of EHMT1, EHMT2, and SUV39H1. To link in vitro catalytic rates with nuclear methylation dynamics, we built a mathematical model incorporating measured kinetic parameters and a time course of mass spectrometry-based H3K9 methylation measurements following cellular SAM depletion. The model revealed that the intrinsic kinetic constants of the catalytic domains could recapitulate in vivo observations. Together, these results suggest catalytic discrimination by H3K9 HMTs maintain nuclear H3K9me1, ensuring epigenetic persistence after metabolic stress.

biochemistry↗

bioRxiv2019

Methyl-Metabolite Depletion Elicits Adaptive Responses to Support Heterochromatin Stability and Epigenetic Persistence

S-adenosylmethionine (SAM) is the methyl-donor substrate for DNA and histone methyltransferases that regulate cellular epigenetic states. This metabolism-epigenome link enables the sensitization of chromatin methylation to altered SAM abundance. However, a chromatin-wide understanding of the adaptive/responsive mechanisms that allow cells to actively protect epigenetic information during life-experienced fluctuations in SAM availability are unknown. We identified a robust response to SAM depletion that is highlighted by preferential cytoplasmic and nuclear de novo mono-methylation of H3 Lys 9 (H3K9) at the expense of global losses in histone di- and tri-methylation. Under SAM-depleted conditions, de novo H3K9 mono-methylation preserves heterochromatin stability and supports global epigenetic persistence upon metabolic recovery. This unique chromatin response was robust across the mouse lifespan and correlated with improved metabolic health, supporting a significant role for epigenetic adaptation to SAM depletion in vivo. Together, these studies provide the first evidence for active epigenetic adaptation and persistence to metabolic stress.

biochemistry↗