Insulin-dependent transcriptional control in L6 rat myotubes is associated with modulation of histone acetylation and accumulation of the histone variant H2A.Z in the proximity of the transcriptional start site

Abstract

Besides its direct metabolic effects, insulin induces transcriptional alterations in its target tissues. However, whether such changes are accompanied by epigenetic changes on the chromatin template encompassing insulin responsive genes is unclear. Here, mRNA levels of insulin-responsive genes hexokinase 2 (Hk2), insulin receptor substrate (Irs2), and the PI3K subunit p85β (Pik3r2) were compared in control versus insulin-stimulated L6 myotubes. Chromatin immunoprecipitation (ChIP) was performed with antibodies directed to histone H2A, histone variant H2A.Z, acetylated histone H3 on lysines 9/14, and acetylated H2A.Z. Insulin induced a more than 2-fold Hk2 mRNA increase, while Irs2 and Pik3r2 were downregulated. ChIP to H2A and H2A.Z showed higher H2A.Z accumulation around the transcriptional start site (TSS) of these insulin-modulated genes, while H2A.Z accumulation was lower distally to the TSS in the Hk2 promoter. H2A.Z levels and H3K9/14 acetylation correlated on several loci along the Hk2 gene, and H3K9/14 as well as H2A.Z acetylation was enhanced by insulin treatment. On the contrary, reduced H3K9/14 acetylation was observed in insulin-repressed Irs2 and Pik3r2, and recovery of acetylation by treatment with the histone deacetylase inhibitor trichostatin A reverted insulin-induced Irs2 downregulation. The chromatin regions encompassing selected insulin-responsive genes are thus featured by accumulation of H2A.Z around the TSS. H2A.Z accumulation facilitates insulin-dependent modulation of pharmacologically treatable H3K9/14 and H2A.Z acetylations. Indeed, inhibition of histone deacetylases by TSA treatment reverted insulin induced Irs2 gene downregulation. Dysregulated histone acetylation may thus be potentially targeted with histone deacetylase inhibitors.

HISTONE DEACETYLASE 9 stimulates auxin-dependent thermomorphogenesis in Arabidopsis thaliana by mediating H2A.Z depletion

“…2D) indicated that ARP6/H2A.Z operates genetically downstream of HDA9. Furthermore, studies in rat (47) and yeast (48) demonstrated that histone acetylation levels positively correlate with H2A.Z occupancy. Taking this information into consideration, we hypothesized that HDA9-mediated H3K9K14 deacetylation at warm temperature conditions could modulate H2A.Z occupancy in Arabidopsis as well, allowing for PIF4 binding followed by thermal induction of YUC8 expression.…”

Prominent action of butyrate over β-hydroxybutyrate as histone deacetylase inhibitor, transcriptional modulator and anti-inflammatory molecule

“…To amplify nuclear and mitochondrial genomic DNA, we used sonicated DNA prepared as previously described 15 . Briefly, cell culture medium was removed and cells were washed twice with ice-cold PBS before cross-linking for 10 minutes with 1% formaldehyde in PBS.…”

Epigenetic modifications in pancreas development, diabetes, and therapeutics

“…Acetylated H2A.Z and H3K9/14 acetylation at the promoter of hexokinase 2 ( HK2 ) increased its expression by twofolds in response to insulin. Similarly, treating rat L6 myotubes with insulin repressed IRS2 and PIK3R2 expression and treatment with trichostatin, a potent and reversible inhibitor of histone deacetylase, resulted in increased histone acetylation at the promoters of Irs2 and Pik3R2 and increased their expression 64 . HDAC6 impairs glucocorticoid‐induced hepatic glucose homeostasis by overexpressing Pepck, G6P and pyruvate carboxylase by changing H4K8 acetylation at the PEPCK gene promoter.…”