Post-translational modifications histone acetylation

As described above, histones are much more than passive structural players within chromatin. Dynamic post-translational modifications of these proteins confer specialized chemical proprieties to chromatin of both informational and structural nature with important functional implications. The highly conserved sites for acetylation, methylation, phosphorylation, ADP-ribosylation, and ubiquitination events on histone tails appear to orchestrate functional activities that range from transcriptional activation and repression to DNA repair and recombination. 

Fig. 6. Post-translational modifications of core and linker histones. The sites of acetylation, phosphorylation, poly-ADP ribosylation, methylation, and ubiquitination are incficated by numbers that correspond to the amino acid position from the N-termini of the molecules. The nomenclature of histone HI variants is as in Fig. 3. The length of C- and N-terminal tails is in relative scale between core histones to illustrate primary structural differences between these proteins.

Experimental results regarding the role of the histone tails indicate that these histone domains play a critical role in chromatin folding [358,365]. Removal as well as the modification (acetylation) of the lysine amino acids within these regions produces an imbalance of the electrostatic interactions, which results in a hierarchically impaired folding ability (H3/H4-H2A/H2B>H3/H4>H2A/H2B) of the chromatin fiber [358,366-369]. Therefore, sources of histone tail variability (histone variants and post-translational modifications other than lysine acetylation) are also likely to alter the extent of folding of chromatin. 

Histone acetylation is without a doubt one of the most thoroughly characterized post-translational modifications of histones where both the functional (see Section 3.1) and structural implications for chromatin have been explored. In the sections that follow we are going to summarize the major structural effects of this post-translational modification as they pertain to the nucleosome and the chromatin fiber. 

A true appreciation of the subtle and complex ways in which the nucleosome can influence gene expression, has come only recently, largely through studies of the post-translational modifications to which all histones are subject and of the enzymes that add and remove these modifications. It has been known for many years that the histone N-terminal tails are exposed on the surface of the nucleosome and that selected amino acid residues are subject to a variety of enzyme-catalyzed, post-translational modifications. These include acetylation of lysines, phosphorylation of serines, and methylation of lysines and arginines ([6,7], see also chapters by Davie, and Ausio and Abbott, this volume). The locations of the histone N-terminal tails in the nucleosome and the residues that can be modified are shown in Fig. 1. 

Fig. 1. Histone modifications on the nucleosome core particle. The nucleosome core particle showing 6 of the 8 core histone N-terminal tail domains and 2 C-terminal tails. Sites of post-translational modification are indicated by coloured symbols that are defined in the key (lower left) acK = acetyl lysine, meR = methyl arginine, mcK = methyl lysine, PS = phosphoryl serine, and uK = ubiquitinated lysine. Residue numbers are shown for each modification. Note that H3 lysine 9 can be either acetylated or methylated. The C-terminal tail domains of one H2A molecule and one H2B molecule are shown (dashed lines) with sites of ubiquitination at H2A lysine 119 (most common in mammals) and H2B lysine 123 (most common in yeast). Modifications are shown on only one of the two copies of histones H3 and H4 and only one tail is shown for H2A and H2B. Sites marked by green arrows are susceptible to cutting by trypsin in intact nucleosomes. Note that the cartoon is a compendium of data from various organisms, some of which may lack particular modifications e.g., there is no H3meK9 in S. cerevisiae. (From Ref [7].)...

In cells of the mammary gland, either in normal epithelial or in cancerous cells, the packaging of chromosomal DNA into chromatin restricts the access of the transcription machinery, thereby causing transcriptional repression. The basic N-termini of histones are subject to post-translational modifications, including lysine acetylation, lysine and arginine methylation, serine phosphorylation and ubiquitinylation [56]. It has been proposed in the histone code hypothesis that the intricate pattern of modifications of the N-terminal histone tail influences gene regulation [57]. 

Overall, histone acetylation and deacetylation represents an important tool with which transcription can be positively or negatively influenced. The nucleosomes and, in a further sense, chromatin structure assiune a central role in the regulation of transcription. Nucleosome structure and nucleosome position can decisively contribute to the accessibility of DNA elements for transcription factors. The nucleosomes function as a framework that determines the spatial arrangement of a region of the DNA. Tlie nucleosome constellation must be modified during transcription initiation, whereby the post-translational modification of histones in the form of acetylation or deacetylation plays a significant role. The participation of other non-histone proteins remains an open issue and it is also imclear how a constitutive and permanent inactivation of a section of DNA can be accomplished via the chromatin structure. 

Histones are basic proteins that are made up by a globular domain and an N-terminal tail that protrudes from the nucleosome. Nucleosomes form the basic unit of chromatin and are made up by a complex of DNA wrapped around an octamer of histones formed by pairs of the histones H2A, H2B, H3, and H4 (45,46) (Fig. 1). Post-translational modification of the core histone tails by methylation, acetylation, phosphorylation, ubiquitina-tion, or sumoylation can alter the structure of the nucleosomes and thus alter gene expression. These post-translational modifications determine the structure and pattern of chromatin condensation and determine the histone code that drives gene transcriptional regulation (47,48). Below are briefly described the factors determining the histone acetylation and methylation. 

Numerous studies have shown that the acetylation or deacetylation of the histones of the nucleosome plays a major role in the regulation of transcriptional activity. Acetylation of the histones (review Narlikar et ah, 2002) is a post-translational modification, which is usually performed on lysine residues at the N-terminus and requires specific enzymes, the histone acetyl transferases (HATs). Removal of the acetyl group also requires specific enzymes, the histone deacetylases [HDAC). Most importantly, the acetylation of histones is accompanied by a loss of positive charges, which is thought to have a profound influence on the nucleosome structure and on the strength of DNA binding. 

Cyclophane-based resorcinarene trimer bearing a dansyl moiety and 28 carboxylate residues Fluorescent probes capable of sensing post-translational modifications of histone Exhibited potent discrimination capability between histone and chemically acetylated histone due to the electrostatic interactions [102]... 

Many proteins rely upon post-translational modification for activity, subcellular localisation or to alter their structure and/or stabiltiy and some of these modifications are reversible. Reversible acetylation for esunple requires the combined action of acetylases (aka acetyltransfoiases) and deacetylases which work n ether to maintain the appropriate acetylation level. It was only just over ten years ago when these enzymes were first identified and in the case of the first deacetylase an inhibitor of the enzyme was used as a probe to purify the protein itself Reversible protein acetylation has been the subject of intense investigation ever since. Although re-seardi has focussed on histone (de)acetylation and its consequences, it is important to consider alternative in vivo substrates. Human histone deacetylase 6 (HDAC6) is a mbulin deacetylase for example (see ref. 2). 

Acetylation and acylation. Acetylation of e-amino groups on conserved Lys residues in the N-terminal domains of the core histones, H2A, H2B, H3 and H4, is the most common post-translational modification of chromatin. Acetylation substantially weakens the constraints on DNA imposed by the core histones and provides molecular mechanism by which DNA becomes accessible to transacting factors while maintaining a nucleosome architecture (Wade et al, 1997). Histone acetylation is reversible and is controlled by a group of acetyltransferases and deacetylases. The balance between histone acetyltransferases and deacetylases determines the accessibility of the chromatin to the transcriptional machinery. Thus the acetylation status of histones is a key determinant of transcriptional activity. Transcription activators are often associated with histone acetyltransferases and repressors can interact with histone deacetylases (Ng and Bird, 2000). 

Histones are core proteins which form nucleosomes with DNA, in which an octamer of two copies of each of four different histones H2A, H2B, H3, and H4 are wrapped by DNA. This nucleosome is a basic unit of chromatin. Post-translational modifications of histones, such as acetylation, methylation, and phosphorylation, play an important role in gene regulation [90]. Although the synthesis of histones H2B [91] and H4 [92, 93], which contain no Cys residues, has been reported based on the combination of NCL and a desulfurization strategy by several groups, the synthesis of the Cys residue-containing histone H3 is difficult and has enjoyed limited success [72, 92, 94]. Recently, one-pot synthesis of H3 using peptide hydrazide as a peptide thioester precursor was reported [95]. 

Wong M, Smulson M (1984) A relationship between nuclear poly (adenosine diphosphate ribosylation) and acetylation post-translational modifications II. Histone Studies Biochemistry 23 3726-3730... 

Figure 5.3 Selected post-translational modifications of human histones. Methylation, acetylation, phosphorylation, biotinylation and ubiquitylation sites have been identified on all four core histones. The modifications shown here should not be regarded as a complete or final list.

The correlation between histone acetylation and eukaryotic transcription were recognized many years ago [128,129]. However, it has not been until very recently, with the discovery that both HATs [130-133] and HDACs [134-138] are an integral part of the basal transcriptional machinery, that the molecular link for this correlation was established. This discovery has rekindled interest in this post-translational histone modification with implications ranging from basic chromatin research to applied medical investigations. Indeed, histone acetylation has been linked to cancer [139-144] and certain types of HDAC inhibitors are already being used to treat certain forms of cancer [145]. 

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