Decreased Histone H4 Acetylation Research Articles

Regulation of nuclear epigenome by mitochondrial DNA heteroplasmy

Diseases associated with mitochondrial DNA (mtDNA) mutations are highly variable in phenotype, in large part because of differences in the percentage of normal and mutant mtDNAs (heteroplasmy) present within the cell. For example, increasing heteroplasmy levels of the mtDNA tRNALeu(UUR) nucleotide (nt) 3243A > G mutation result successively in diabetes, neuromuscular degenerative disease, and perinatal lethality. These phenotypes are associated with differences in mitochondrial function and nuclear DNA (nDNA) gene expression, which are recapitulated in cybrid cell lines with different percentages of m.3243G mutant mtDNAs. Using metabolic tracing, histone mass spectrometry, and NADH fluorescence lifetime imaging microscopy in these cells, we now show that increasing levels of this single mtDNA mutation cause profound changes in the nuclear epigenome. At high heteroplasmy, mitochondrially derived acetyl-CoA levels decrease causing decreased histone H4 acetylation, with glutamine-derived acetyl-CoA compensating when glucose-derived acetyl-CoA is limiting. In contrast, α-ketoglutarate levels increase at midlevel heteroplasmy and are inversely correlated with histone H3 methylation. Inhibition of mitochondrial protein synthesis induces acetylation and methylation changes, and restoration of mitochondrial function reverses these effects. mtDNA heteroplasmy also affects mitochondrial NAD+/NADH ratio, which correlates with nuclear histone acetylation, whereas nuclear NAD+/NADH ratio correlates with changes in nDNA and mtDNA transcription. Thus, mutations in the mtDNA cause distinct metabolic and epigenomic changes at different heteroplasmy levels, potentially explaining transcriptional and phenotypic variability of mitochondrial disease.

MC1568 Inhibits Thimerosal-Induced Apoptotic Cell Death by Preventing HDAC4 Up-Regulation in Neuronal Cells and in Rat Prefrontal Cortex.

Ethylmercury thiosalicylate (thimerosal) is an organic mercury-based compound commonly used as an antimicrobial preservative that has been found to be neurotoxic. In contrast, histone deacetylases (HDACs) inhibition has been found to be neuroprotective against several environmental contaminants, such as polychlorinated biphenyls, di-2-ethylhexyl phthalate, and methylmercury. The aim of this study was to investigate the effect of HDAC inhibition on thimerosal-induced neurotoxicity in neuroblastoma cells and cortical neurons. Interestingly, we found that thimerosal, at 0.5 μM in SH-SY5Y cells and at 1 μM in neurons, caused cell death by activation of apoptosis, which was prevented by the HDAC class IIA inhibitor MC1568 but not the class I inhibitor MS275. Furthermore, thimerosal specifically increased HDAC4 protein expression but not that of HDACs 5, 6, 7, and 9. Western blot analysis revealed that MC1568 prevented thimerosal-induced HDAC4 increase. In addition, both HDAC4 knocking-down and MC1568 inhibited thimerosal-induced cell death in SH-SY5Y cells and cortical neurons. Importantly, intramuscular injection of 12 μg/kg thimerosal on postnatal days 7, 9, 11, and 15 increased HDAC4 levels in the prefrontal cortex (PFC), which decreased histone H4 acetylation in infant male rats, in parallel increased motor activity changes. In addition, coadministration of 40 mg/kg MC1568 (intraperitoneal injection) moderated the HDAC4 increase which reduced histone H4 deacetylation and caspase-3 cleavage in the PFC. Finally, open-field testing showed that thimerosal-induced motor activity changes are reduced by MC1568. These findings indicate that HDAC4 regulates thimerosal-induced cell death in neurons and that treatment with MC1568 prevents thimerosal-induced activation of caspase-3 in the rat PFC.

Expression and functional analysis of the plant-specific histone deacetylase HDT701 in rice

Reversible histone acetylation and deacetylation at the N-terminus of histone tails play a crucial role in regulating eukaryotic gene activity. Acetylation of core histones is associated with gene activation, whereas deacetylation of histone is often correlated with gene repression. The level of histone acetylation is antagonistically catalyzed by histone acetyltransferases citation(HATs) and histone deacetylases (HDACs). In this work, we examined the subcellular localization, expression pattern and function of HDT701, a member of the plant-specific HD2-type histone deacetylase in rice. HDT701 is localized at the subcellular level in the nucleus. Histochemical GUS-staining analysis revealed that HDT701 is constitutively expressed throughout the life cycle of rice. Overexpression of HDT701 in rice decreases ABA, salt and osmotic stress resistance during seed germination. Delayed seed germination of HDT701 overexpression lines is associated with decreased histone H4 acetylation and down-regulated expression of GA biosynthetic genes. Moreover, overexpression of HDT701 in rice enhances salt and osmotic stress resistance during the seedling stage. Taken together, our findings suggested that HDT701 may play an important role in regulating seed germination in response to abiotic stresses in rice.

Fe65 is required for Tip60-directed histone H4 acetylation at DNA strand breaks

Fe65 is a binding partner of the Alzheimer's beta-amyloid precursor protein APP. The possible involvement of this protein in the cellular response to DNA damage was suggested by the observation that Fe65 null mice are more sensitive to genotoxic stress than WT counterpart. Fe65 associated with chromatin under basal conditions and its involvement in DNA damage repair requires this association. A known partner of Fe65 is the histone acetyltransferase Tip60. Considering the crucial role of Tip60 in DNA repair, we explored the hypothesis that the phenotype of Fe65 null cells depended on its interaction with Tip60. We demonstrated that Fe65 knockdown impaired recruitment of Tip60-TRRAP complex to DNA double strand breaks and decreased histone H4 acetylation. Accordingly, the efficiency of DNA repair was decreased upon Fe65 suppression. To explore whether APP has a role in this mechanism, we analyzed a Fe65 mutant unable to bind to APP. This mutant failed to rescue the phenotypes of Fe65 null cells; furthermore, APP/APLP2 suppression results in the impairment of recruitment of Tip60-TRRAP complex to DNA double strand breaks, decreased histone H4 acetylation and repair efficiency. On these bases, we propose that Fe65 and its interaction with APP play an important role in the response to DNA damage by assisting the recruitment of Tip60-TRRAP to DNA damage sites.

Atherosclerosis-prone hemodynamics differentially regulates endothelial and smooth muscle cell phenotypes and promotes pro-inflammatory priming

Atherosclerosis is an inflammatory disease that preferentially forms at hemodynamically compromised regions of altered shear stress patterns. Endothelial cells (EC) and smooth muscle cells (SMC) undergo phenotypic modulation during atherosclerosis. An in vitro coculture model was developed to determine the role of hemodynamic regulation of EC and SMC phenotypes in coculture. Human ECs and SMCs were plated on a synthetic elastic lamina and human-derived atheroprone, and atheroprotective shear stresses were imposed on ECs. Atheroprone flow decreased genes associated with differentiated ECs (endothelial nitric oxide synthase, Tie2, and Kruppel-like factor 2) and SMCs (smooth muscle alpha-actin and myocardin) and induced a proinflammatory phenotype in ECs and SMCs (VCAM-1, IL-8, and monocyte chemoattractant protein-1). Atheroprone flow-induced changes in SMC differentiation markers were regulated at the chromatin level, as indicated by decreased serum response factor (SRF) binding to the smooth muscle alpha-actin-CC(a/T)(6)GG (CArG) promoter region and decreased histone H(4) acetylation. Conversely, SRF and histone H(4) acetylation were enriched at the c-fos promoter in SMCs. In the presence of atheroprotective shear stresses, ECs aligned with the direction of flow and SMCs aligned more perpendicular to flow, similar to in vivo vessel organization. These results provide a novel mechanism whereby modulation of the EC phenotype by hemodynamic shear stresses, atheroprone or atheroprotective, play a critical role in mechanical-transcriptional coupling and regulation of the SMC phenotype.

Glucose regulation of insulin gene expression requires the recruitment of p300 by the beta-cell-specific transcription factor Pdx-1.

Regulation of insulin gene expression in response to increases in blood glucose levels is essential for maintaining normal glucose homeostasis; however, the exact mechanisms by which glucose stimulates insulin gene transcription are not known. We have shown previously that glucose stimulates insulin gene expression by causing the hyperacetylation of histone H4 at the insulin promoter. We demonstrate that the histone acetyltransferase p300 is recruited to the insulin promoter only at high concentrations of glucose via its interaction with the beta-cell-specific transcription factor Pdx-1. Disruption of the function of the endogenous Pdx-1 abolishes the recruitment of p300 to the insulin gene promoter at high concentrations of glucose and results in decreased histone H4 acetylation and insulin gene expression. Furthermore, we demonstrate that the glucose-dependent interaction of Pdx-1 with p300 is regulated by a phosphorylation event that changes the localization of Pdx-1. Based on these data, we conclude that hyperacetylation of histone H4 at the insulin gene promoter in response to high concentrations of glucose depends on the beta-cell-specific transcription factor Pdx-1, which is required for the recruitment of the histone acetyltransferase p300 to the insulin gene promoter.