Host cell factor and an uncharacterized SANT domain protein are stable components of ATAC, a novel dAda2A/dGcn5-containing histone acetyltransferase complex in Drosophila

Gcn5 is a conserved histone acetyltransferase (HAT) found in a number of multisubunit complexes from Saccharomyces cerevisiae, mammals, and flies. We previously identified Drosophila melanogaster homologues of the yeast proteins Ada2, Ada3, Spt3, and Tra1 and showed that they associate with dGcn5 to form at least two distinct HAT complexes. There are two different Ada2 homologues in Drosophila named dAda2A and dAda2B. dAda2B functions within the Drosophila version of the SAGA complex (dSAGA). To gain insight into dAda2A function, we sought to identify novel components of the complex containing this protein, ATAC (Ada two A containing) complex. Affinity purification and mass spectrometry revealed that, in addition to dAda3 and dGcn5, host cell factor (dHCF) and a novel SANT domain protein, named Atac1 (ATAC component 1), copurify with this complex. Coimmunoprecipitation experiments confirmed that these proteins associate with dGcn5 and dAda2A, but not with dSAGA-specific components such as dAda2B and dSpt3. Biochemical fractionation revealed that ATAC has an apparent molecular mass of 700 kDa and contains dAda2A, dGcn5, dAda3, dHCF, and Atac1 as stable subunits. Thus, ATAC represents a novel histone acetyltransferase complex that is distinct from previously purified Gcn5/Pcaf-containing complexes from yeast and mammalian cells.     

Epigenetic regulation of nuclear lamina-associated heterochromatin by HAT1 and the acetylation of newly synthesized histones

A central component of the epigenome is the pattern of histone post-translational modifications that play a critical role in the formation of specific chromatin states. Following DNA replication, nascent chromatin is a 1:1 mixture of parental and newly synthesized histones and the transfer of modification patterns from parental histones to new histones is a fundamental step in epigenetic inheritance. Here we report that loss of HAT1, which acetylates lysines 5 and 12 of newly synthesized histone H4 during replication-coupled chromatin assembly, results in the loss of accessibility of large domains of heterochromatin, termed HAT1-dependent Accessibility Domains (HADs). HADs are mega base-scale domains that comprise ∼10% of the mouse genome. HAT1 globally represses H3 K9 me3 levels and HADs correspond to the regions of the genome that display HAT1-dependent increases in H3 K9me3 peak density. HADs display a high degree of overlap with a subset of Lamin-Associated Domains (LADs). HAT1 is required to maintain nuclear structure and integrity. These results indicate that HAT1 and the acetylation of newly synthesized histones may be critical regulators of the epigenetic inheritance of heterochromatin and suggest a new mechanism for the epigenetic regulation of nuclear lamina-heterochromatin interactions.     

Acetylation of Rsc4p by Gcn5p is essential in the absence of histone H3 acetylation

Rsc4p, a subunit of the RSC chromatin-remodeling complex, is acetylated at lysine 25 by Gcn5p, a well-characterized histone acetyltransferase (HAT). Mutation of lysine 25 does not result in a significant growth defect, and therefore whether this modification is important for the function of the essential RSC complex was unknown. In a search to uncover the molecular basis for the lethality resulting from loss of multiple histone H3-specific HATs, we determined that loss of Rsc4p acetylation is lethal in strains lacking histone H3 acetylation. Phenotype comparison of mutants with arginine and glutamine substitutions of acetylatable lysines within the histone H3 tail suggests that it is a failure to neutralize the charge of the H3 tail that is lethal in strains lacking Rsc4p acetylation. We also demonstrate that Rsc4p acetylation does not require any of the known Gcn5p-dependent HAT complexes and thus represents a truly novel function for Gcn5p. These results demonstrate for the first time the vital and yet redundant functions of histone H3 and Rsc4p acetylation in maintaining cell viability.     

Glucocorticoid receptor recruitment of histone deacetylase 2 inhibits interleukin-1beta-induced histone H4 acetylation on lysines 8 and 12

We have investigated the ability of dexamethasone to regulate interleukin-1beta (IL-1beta)-induced gene expression, histone acetyltransferase (HAT) and histone deacetylase (HDAC) activity. Low concentrations of dexamethasone (10(-10) M) repress IL-1beta-stimulated granulocyte-macrophage colony-stimulating factor (GM-CSF) expression and fail to stimulate secretory leukocyte proteinase inhibitor expression. Dexamethasone (10(-7) M) and IL-1beta (1 ng/ml) both stimulated HAT activity but showed a different pattern of histone H4 acetylation. Dexamethasone targeted lysines K5 and K16, whereas IL-1beta targeted K8 and K12. Low concentrations of dexamethasone (10(-10) M), which do not transactivate, repressed IL-1beta-stimulated K8 and K12 acetylation. Using chromatin immunoprecipitation assays, we show that dexamethasone inhibits IL-1beta-enhanced acetylated K8-associated GM-CSF promoter enrichment in a concentration-dependent manner. Neither IL-1beta nor dexamethasone elicited any GM-CSF promoter association at acetylated K5 residues. Furthermore, we show that GR acts both as a direct inhibitor of CREB binding protein (CBP)-associated HAT activity and also by recruiting HDAC2 to the p65-CBP HAT complex. This action does not involve de novo synthesis of HDAC protein or altered expression of CBP or p300/CBP-associated factor. This mechanism for glucocorticoid repression is novel and establishes that inhibition of histone acetylation is an additional level of control of inflammatory gene expression. This further suggests that pharmacological manipulation of of specific histone acetylation status is a potentially useful approach for the treatment of inflammatory diseases.     

Arabidopsis GCN5, HD1, and TAF1/HAF2 interact to regulate histone acetylation required for light-responsive gene expression

We previously showed that Arabidopsis thaliana histone acetyltransferase TAF1/HAF2 is required for the light regulation of growth and gene expression, and we show here that histone acetyltransferase GCN5 and histone deacetylase HD1/HDA19 are also involved in such regulation. Mutation of GCN5 resulted in a long-hypocotyl phenotype and reduced light-inducible gene expression, whereas mutation of HD1 induced opposite effects. The double mutant gcn5 hd1 restored a normal photomorphogenic phenotype. By contrast, the double mutant gcn5 taf1 resulted in further loss of light-regulated gene expression. gcn5 reduced acetylation of histones H3 and H4, mostly on the core promoter regions, whereas hd1 increased acetylation on both core and more upstream promoter regions. GCN5 and TAF1 were both required for H3K9, H3K27, and H4K12 acetylation on the target promoters, but H3K14 acetylation was dependent only on GCN5. Interestingly, gcn5 taf1 had a cumulative effect mainly on H3K9 acetylation. On the other hand, hd1 induced increased acetylation on H3K9, H3K27, H4K5, and H4K8. GCN5 was also shown to be directly associated with the light-responsive promoters. These results suggest that acetylation of specific histone Lys residues, regulated by GCN5, TAF1, and HD1, is required for light-regulated gene expression.     

Highly specific antibodies determine histone acetylation site usage in yeast heterochromatin and euchromatin.

We have developed a highly specific antibody set for acetylation sites in yeast histones H4 (K5, K8, K12, and K16); H3 (K9, K14, K18, K23, and K27); H2A (K7); and H2B (K11 and K16). Since ELISA does not assure antibody specificity in chromatin immunoprecipitation, we have employed additional screens against the respective histone mutations. We now show that telomeric and silent mating locus heterochromatin is hypoacetylated at all histone sites. At the INO1 promoter, RPD3 is required for strongly deacetylating all sites except H4 K16, ESA1 for acetylating H2A, H2B, and H4 sites except H4 K16, and GCN5 for acetylating H2B and H3 sites except H3 K14. These data uncover the in vivo usage of acetylation sites in heterochromatin and euchromatin.     

Solar Simulated Ultraviolet Radiation Induces Global Histone Hypoacetylation in Human Keratinocytes

Ultraviolet radiation (UVR) from sunlight is the primary effector of skin DNA damage. Chromatin remodeling and histone post-translational modification (PTM) are critical factors in repairing DNA damage and maintaining genomic integrity, however, the dynamic changes of histone marks in response to solar UVR are not well characterized. Here we report global changes in histone PTMs induced by solar simulated UVR (ssUVR). A decrease in lysine acetylation of histones H3 and H4, particularly at positions of H3 lysine 9, lysine 56, H4 lysine 5, and lysine 16, was found in human keratinocytes exposed to ssUVR. These acetylation changes were highly associated with ssUVR in a dose-dependent and time-specific manner. Interestingly, H4K16ac, a mark that is crucial for higher order chromatin structure, exhibited a persistent reduction by ssUVR that was transmitted through multiple cell divisions. In addition, the enzymatic activities of histone acetyltransferases were significantly reduced in irradiated cells, which may account for decreased global acetylation. Moreover, depletion of histone deacetylase SIRT1 in keratinocytes rescued ssUVR-induced H4K16 hypoacetylation. These results indicate that ssUVR affects both HDAC and HAT activities, leading to reduced histone acetylation.