Acetylation increases the alpha-helical content of the histone tails of the nucleosome

The nature of the structural changes induced by histone acetylation at the different levels of chromatin organization has been very elusive. At the histone level, it has been proposed on several occasions that acetylation may induce an alpha-helical conformation of their acetylated N-terminal domains (tails). In an attempt to provide experimental support for this hypothesis, we have purified and characterized the tail of histone H4 in its native and mono-, di-, tri-, and tetra- acetylated form. The circular dichroism analysis of these peptides shows conclusively that acetylation does increase their alpha-helical content. Furthermore, the same spectroscopic analysis shows that this is also true for both the acetylated nucleosome core particle and the whole histone octamer in solution. In contrast to the native tails in which the alpha-helical organization appears to be dependent upon interaction of these histone regions with DNA, the acetylated tails show an increase in alpha-helical content that does not depend on such an interaction.     

Biochemical characterization of chromatin fractions isolated from induced and uninduced friend erythroleukemia cells

Summary Chromatin fractions from Friend erythroleukemia cells after induction of differentiation by dimethylsulfoxide (DMSO) were compared in their biochemical characteristics to fractions from uninduced cells. Fractions were prepared by extracting chromatin from nuclei after mild micrococcal nuclease treatment with increasing concentrations of NaCl according to Sanders [1]. This procedure has been found to release chromatin containing hyperacetylated histones preferentially [2]. The fractions obtained by this procedure were analysed in respect to the amount of chromatin released, the amount of histone H1, the degree of acetylation of histone H4, the presence of non-histone proteins and the concentration of transcribed and non-transcribed sequences. It was found that the fractions differ in the amount of histone H1 present, in several non-histone proteins and in the acetylation of histonie H4, regardless whether induced or uninduced cells were analysed. The distribution of transcribed sequences versus non-transcribed sequences among the fractions was the same, demonstrating that this fractionation procedure, although leading to fractions with biochemical differences, is not able to discriminate functional states of chromatin and that the biochemical characteristics of the fractions may be common to both, active as well as inactive states of chromatin.     

Histone acetylation and chromatin pattern in cancer. A review

Chromatin phenotype is known to be significantly disrupted in cancer. This has been demonstrated in many morphologic studies on cancer and in recent years by the application of digital texture analysis for quantitative evaluation of chromatin phenotype in neoplasia. Studies have consistently demonstrated the role of chromatin phenotype as a biomarker of diagnosis and prognosis. The underlying molecular mechanisms for chromatin reorganization and its role as a biomarker are largely unknown, but epigenetic processes are likely to be a main factor that not only modify chromatin arrangement but in doing so alter gene expression profiles in a reversible fashion. Of the range of epigenetic modifications that might control chromatin phenotype, histone acetylation is a strong candidate because of its role in the direct modification of chromatin, both through local relaxation of nucleosomal structure and recruitment of chromatin remodeling complexes. The reversible nature of histone acetylation is therapeutically attractive for treatment of aberrant histone acetylation; however, it still remains to be seen whether histone deacetylase inhibitors are clinically applicable or for use primarily as valuable research tools. This review explores the role of histone acetylation in cancer development, as a potential therapeutic candidate and a potential biomarker in tissue pathology.     

Methods and tips for the purification of human histone methyltransferases

Recently developed biochemical techniques have enabled researchers to study histone modifications more easily and accurately. One of these modifications, histone lysine methylation, has been shown to be highly stable and to represent an epigenetic alteration. Extensive biochemical analyses have led to discoveries about the nature and functions of this modification, thus accelerating our understanding of this crucial epigenetic event. Here we describe basic methods for purification and biochemical analysis of lysine-directed, histone methyltransferases from HeLa cell-derived extracts. In the section on substrate preparation, we describe a simple method for the preparation of recombinant substrates, although we recommend using native substrates for initial detection of the activities. The purification protocols for several histone methyltransferases have been streamlined so that those researchers with a basic understanding of biochemistry can perform them. We also describe many tips and provide suggestions to avoid common pitfalls in the biochemical analysis of histone methyltransferases.     

Histone acetylation during early stages of sea urchin (arbacia punctulata) development

A correlation has been found between histone acetylation and gene activation at the time of gastrulation in the sea urchin, Arbacia punctulata. Between the blastula and gastrula stages, there is a 2.5-fold increase in the rate of acetylation of a histone fraction consisting of slightly lysine-rich and argininerich components, with a marked decrease in the fully differentiated pluteus stage. This pre-gastrular increase in histone acetylation is not correlated with (1) a decrease in the rate of histone deacetylation; (2) a decrease in acetyl coenzyme A pool size; (3) an increase in acetate uptake; (4) histone synthesis. The results thus suggest that increased histone acetylation may be at least one preparative factor for the activation of new genes at gastrulation.     

Role of Jade-1 in the histone acetyltransferase (HAT) HBO1 complex

Regulation of global chromatin acetylation is important for chromatin remodeling. A small family of Jade proteins includes Jade-1L, Jade-2, and Jade-3, each bearing two mid-molecule tandem plant homology domain (PHD) zinc fingers. We previously demonstrated that the short isoform of Jade-1L protein, Jade-1, is associated with endogenous histone acetyltransferase (HAT) activity. It has been found that Jade-1L/2/3 proteins co-purify with a novel HAT complex, consisting of HBO1, ING4/5, and Eaf6. We investigated a role for Jade-1/1L in the HBO1 complex. When overexpressed individually, neither Jade-1/1L nor HBO1 affected histone acetylation. However, co-expression of Jade-1/1L and HBO1 increased acetylation of the bulk of endogenous histone H4 in epithelial cells in a synergistic manner, suggesting that Jade1/1L positively regulates HBO1 HAT activity. Conversely, small interfering RNA-mediated depletion of endogenous Jade resulted in reduced levels of H4 acetylation. Moreover, HBO1-mediated H4 acetylation activity was enhanced severalfold by the presence of Jade-1/1L in vitro. The removal of PHD fingers affected neither binding nor mutual Jade-1-HBO1 stabilization but completely abrogated the synergistic Jade-1/1L- and HBO1-mediated histone H4 acetylation in live cells and in vitro with reconstituted oligonucleosome substrates. Therefore, PHDs are necessary for Jade-1/1L-induced acetylation of nucleosomal histones by HBO1. In contrast to Jade-1/1L, the PHD zinc finger protein ING4/5 failed to synergize with HBO1 to promote histone acetylation. The physical interaction of ING4/5 with HBO1 occurred in the presence of Jade-1L or Jade-3 but not with the Jade-1 short isoform. In summary, this study demonstrates that Jade-1/1L are crucial co-factors for HBO1-mediated histone H4 acetylation.     

Sites of Acetylation on Newly Synthesized Histone H4 Are Required for Chromatin Assembly and DNA Damage Response Signaling

The best-characterized acetylation of newly synthesized histone H4 is the diacetylation of the NH₂-terminal tail on lysines 5 and 12. Despite its evolutionary conservation, this pattern of modification has not been shown to be essential for either viability or chromatin assembly in any model organism. We demonstrate that mutations in histone H4 lysines 5 and 12 in yeast confer hypersensitivity to replication stress and DNA-damaging agents when combined with mutations in histone H4 lysine 91, which has also been found to be a site of acetylation on soluble histone H4. In addition, these mutations confer a dramatic decrease in cell viability when combined with mutations in histone H3 lysine 56. We also show that mutation of the sites of acetylation on newly synthesized histone H4 results in defects in the reassembly of chromatin structure that accompanies the repair of HO-mediated double-strand breaks. This defect is not due to a decrease in the level of histone H3 lysine 56 acetylation. Intriguingly, mutations that alter the sites of newly synthesized histone H4 acetylation display a marked decrease in levels of phosphorylated H2A (γ-H2AX) in chromatin surrounding the double-strand break. These results indicate that the sites of acetylation on newly synthesized histones H3 and H4 can function in nonoverlapping ways that are required for chromatin assembly, viability, and DNA damage response signaling.     

Butyrate-induced GPR41 activation inhibits histone acetylation and cell growth

Butyrate has been recently identified as a natural ligand of the G-protein-coupled receptor 41(GPR41).In addition,it is an inhibitor of histone deacetylase(HDAC).Butyrate treatment results in the hyperacetylation of histones,with resultant multiple biological effects including inhibition of proliferation,induction of cell cycle arrest,and apoptosis,in a variety of cultured mammalian cells.However,it is not clear whether GPR41 is actively involved in the above-mentioned processes.In this study,we generated a stable cell line expressing the hGPR41 receptor in order to investigate the involvement of GPR41 on butyrate-induced biochemical and physiologic processes.We found that GPR41 activation may be a compensatory mechanism to counter the increase in histone H3 acetylation levels induced by butyrate treatment.Moreover,GPR41 had an inhibitory effect on the anti-proliferative,pro-apoptotic effects of butyrate.GPR41 expression induced cell cycle arrest at the G1-stage,while its activation by butyrate can cause more cells to pass the G1 checkpoint.These results indicated that GPR41 was associated with histone acetylation and might be involved in the acetylation-related regulation of cell processes including proliferation,apoptosis,and the cell cycle.