Dynamic acetylation of all lysine 4-methylated histone H3 in the mouse nucleus: analysis at c-fos and c-jun

A major focus of current research into gene induction relates to chromatin and nucleosomal regulation, especially the significance of multiple histone modifications such as phosphorylation, acetylation, and methylation during this process. We have discovered a novel physiological characteristic of all lysine 4 (K4)–methylated histone H3 in the mouse nucleus, distinguishing it from lysine 9–methylated H3. K4-methylated histone H3 is subject to continuous dynamic turnover of acetylation, whereas lysine 9–methylated H3 is not. We have previously reported dynamic histone H3 phosphorylation and acetylation as a key characteristic of the inducible proto-oncogenes c-fos and c-jun. We show here that dynamically acetylated histone H3 at these genes is also K4-methylated. Although all three modifications are proven to co-exist on the same nucleosome at these genes, phosphorylation and acetylation appear transiently during gene induction, whereas K4 methylation remains detectable throughout this process. Finally, we address the functional significance of the turnover of histone acetylation on the process of gene induction. We find that inhibition of turnover, despite causing enhanced histone acetylation at these genes, produces immediate inhibition of gene induction. These data show that all K4-methylated histone H3 is subject to the continuous action of HATs and HDACs, and indicates that at c-fos and c-jun, contrary to the predominant model, turnover and not stably enhanced acetylation is relevant for efficient gene induction.     

Dynamic Acetylation of All Lysine 4–Methylated Histone H3 in the Mouse Nucleus: Analysis at c-fos and c-jun

A major focus of current research into gene induction relates to chromatin and nucleosomal regulation, especially the significance of multiple histone modifications such as phosphorylation, acetylation, and methylation during this process. We have discovered a novel physiological characteristic of all lysine 4 (K4)–methylated histone H3 in the mouse nucleus, distinguishing it from lysine 9–methylated H3. K4-methylated histone H3 is subject to continuous dynamic turnover of acetylation, whereas lysine 9–methylated H3 is not. We have previously reported dynamic histone H3 phosphorylation and acetylation as a key characteristic of the inducible proto-oncogenes c-fos and c-jun. We show here that dynamically acetylated histone H3 at these genes is also K4-methylated. Although all three modifications are proven to co-exist on the same nucleosome at these genes, phosphorylation and acetylation appear transiently during gene induction, whereas K4 methylation remains detectable throughout this process. Finally, we address the functional significance of the turnover of histone acetylation on the process of gene induction. We find that inhibition of turnover, despite causing enhanced histone acetylation at these genes, produces immediate inhibition of gene induction. These data show that all K4-methylated histone H3 is subject to the continuous action of HATs and HDACs, and indicates that at c-fos and c-jun, contrary to the predominant model, turnover and not stably enhanced acetylation is relevant for efficient gene induction.     

K562 human erythroleukemia cell variants resistant to growth inhibition by butyrate have deficient histone acetylation

K562 is an established human erythroleukemia cell line, inducible for hemoglobin synthesis by a variety of compounds including n-butyrate. To elucidate the role of butyrate-induced histone acetylation in the regulation of gene expression in K562 cells, we isolated 20 variants resistant to the growth inhibitory effect of butyrate. Four variants having different degrees of resistance were selected for detailed study. All four were found to be resistant to the hemoglobin-inducing effect of butyrate, suggesting that the two aspects of butyrate response, restriction of growth and induction of hemoglobin synthesis, are coupled. Further, after (5 days) culture with butyrate, two of the four variants exhibit less acetylation of H3 and H4 histones than does the butyrate-treated parent. Analysis of histone deacetylases from the variants indicated that each variant was distinct and that butyrate resistance may be accounted for by decreased affinity of the variant enzymes for butyrate, increased affinity of the enzymes for acetylated histone, or both. The fact that variants selected for resistance to growth inhibition by butyrate are also deficient in butyrate-induced hemoglobin synthesis and have abnormal histone deacetylase activity argues for butyrate inducing K562 cells to synthesize hemoglobin and restrict growth via histone acetylation.     

Histone deacetylase inhibitors induce apoptosis in peripheral blood lymphocytes along with histone H4 acetylation and the expression of the linker histone variant, H1 degrees

The results of this study show that H1 degrees can be induced by sodium butyrate and trichostatin A in peripheral blood lymphocytes, a cell system which does not normally express this linker histone variant. Moreover, this induced expression was found to be correlated in a dose-dependent manner with the concomitant induction of apoptosis and increased levels of histone H4 acetylation. Sodium butyrate and trichostatin A, both inhibitors of histone deacetylases, are known to induce terminal differentiation and at the same time the induction of the linker histone variant, H1 degrees, in a number of tissue/cell systems. Moreover, aside from induced expression by histone deacetylase inhibitors, H1 degrees gene expression has also been tightly associated with the process of terminal differentiation in many physiological tissue/cell systems. The concomitant induction of H1 degrees expression along with apoptosis and histone acetylation in the same cell system has not been previously reported. Histone acetylation is known to be involved in chromatin remodelling events. Such events also occur during apoptosis. The association of H1 degrees gene expression with apoptosis, and not with differentiation in these cells, leads to more general implications as to a potential functional role of H1 degrees during chromatin remodelling.     

Organismal differences in post-translational modifications in histones H3 and H4

Post-translational modifications (PTMs) of histones play an important role in many cellular processes, notably gene regulation. Using a combination of mass spectrometric and immunobiochemical approaches, we show that the PTM profile of histone H3 differs significantly among the various model organisms examined. Unicellular eukaryotes, such as Saccharomyces cerevisiae (yeast) and Tetrahymena thermophila (Tet), for example, contain more activation than silencing marks as compared with mammalian cells (mouse and human), which are generally enriched in PTMs more often associated with gene silencing. Close examination reveals that many of the better-known modified lysines (Lys) can be either methylated or acetylated and that the overall modification patterns become more complex from unicellular eukaryotes to mammals. Additionally, novel species-specific H3 PTMs from wild-type asynchronously grown cells are also detected by mass spectrometry. Our results suggest that some PTMs are more conserved than previously thought, including H3K9me1 and H4K20me2 in yeast and H3K27me1, -me2, and -me3 in Tet. On histone H4, methylation at Lys-20 showed a similar pattern as H3 methylation at Lys-9, with mammals containing more methylation than the unicellular organisms. Additionally, modification profiles of H4 acetylation were very similar among the organisms examined.     

Cross-talk between histone modifications in response to histone deacetylase inhibitors: MLL4 links histone H3 acetylation and histone H3K4 methylation

Histones are subject to a wide variety of post-translational modifications that play a central role in gene activation and silencing. We have used histone modification-specific antibodies to demonstrate that two histone modifications involved in gene activation, histone H3 acetylation and H3 lysine 4 methylation, are functionally linked. This interaction, in which the extent of histone H3 acetylation determines both the abundance and the "degree" of H3K4 methylation, plays a major role in the epigenetic response to histone deacetylase inhibitors. A combination of in vivo knockdown experiments and in vitro methyltransferase assays shows that the abundance of H3K4 methylation is regulated by the activities of two opposing enzyme activities, the methyltransferase MLL4, which is stimulated by acetylated substrates, and a novel and as yet unidentified H3K4me3 demethylase.     

Induction by Fructose Force-Feeding of Histone H3 and H4 Acetylation at Their Lysine Residues around the Slc2a5 Gene and Its Expression in Mice

It has been reported that fructose force-feeding rapidly induced jejunal Slc2a5 gene expression in rodents. We demonstrate in this study that acetylation at lysine (K) 9 of histone H3 and acetylation at K5 and K16 of histone H4 were more enhanced in the promoter/enhancer to transcribed regions of the Slc2a5 gene in fructose force-fed mice than in glucose force-fed mice. However, fructose force-feeding did not induce acetylation at K14 of histone H3, or at K8 and K12 of histone H4 around the Slc2a5 gene. These results suggest that fructose force-feeding induced selective histone acetylation, particularly of H3 and H4, around the jejunal Slc2a5 gene in mice.     

Overlapping Functions between SWR1 Deletion and H3K56 Acetylation in Candida albicans

Nucleosome destabilization by histone variants and modifications has been implicated in the epigenetic regulation of gene expression, with the histone variant H2A.Z and acetylation of H3K56 (H3K56ac) being two examples. Here we find that deletion of SWR1, the major subunit of the SWR1 complex depositing H2A.Z into chromatin in exchange for H2A, promotes epigenetic white-opaque switching in Candida albicans. We demonstrate through nucleosome mapping that SWR1 is required for proper nucleosome positioning on the promoter of WOR1, the master regulator of switching, and that its effects differ in white and opaque cells. Furthermore, we find that H2A.Z is enriched adjacent to nucleosome-free regions at the WOR1 promoter in white cells, suggesting a role in the stabilization of a repressive chromatin state. Deletion of YNG2, a subunit of the NuA4 H4 histone acetyltransferase (HAT) that targets SWR1 activity through histone acetylation, produces a switching phenotype similar to that of swr1, and both may act downstream of the GlcNAc signaling pathway. We further uncovered a genetic interaction between swr1 and elevated H3K56ac with the discovery that the swr1 deletion mutant is highly sensitive to nicotinamide. Our results suggest that the interaction of H2A.Z and H3K56ac regulates epigenetic switching at the nucleosome level, as well as having global effects.