Abstract
BackgroundThe involvement of histone acetylation in facilitating gene expression is well-established, particularly in the case of histones H3 and H4. It was previously shown in Saccharomyces cerevisiae that gene expression was significantly down-regulated and chromatin more condensed in stationary phase compared to exponential phase. We were therefore interested in establishing the acetylation state of histone H3 and H4 in stationary and in exponential phase, since the regulation of this modification could contribute to transcriptional shut-down and chromatin compaction during semi-quiescence.ResultsWe made use of nano-spray tandem mass spectrometry to perform a precursor ion scan to detect an m/z 126 immonium ion, diagnostic of an Nε-acetylated lysine residue that allowed unambiguous identification of acetylated as opposed to tri-methylated lysine. The fragmentation spectra of peptides thus identified were searched with Mascot against the Swiss-Prot database, and the y-ion and b-ion fragmentation series subsequently analyzed for mass shifts compatible with acetylated lysine residues. We found that K9, K14 and K36 of histone H3 and K12 and K16 of histone H4 were acetylated in exponential phase (bulk histones), but could not detect these modifications in histones isolated from stationary phase cells at the sensitivity level of the mass spectrometer. The corresponding un-acetylated peptides were, however, observed. A significantly higher level of acetylation of these residues in exponential phase was confirmed by immuno-blotting.ConclusionH4K16 acetylation was previously shown to disrupt formation of condensed chromatin in vitro. We propose that de-acetylation of H4K16 allowed formation of condensed chromatin in stationary phase, and that acetylation of H3K9, H3K14, H3K36, and H4K12 reflected the active transcriptional state of the yeast genome in exponential phase.
Highlights
The involvement of histone acetylation in facilitating gene expression is well-established, in the case of histones H3 and H4
We have previously shown that linker histone Hho1 displayed a significant increase in chromatin binding in stationary phase in Saccharomyces cerevisiae, which coincided with an increase in chromatin compaction [8]
We were interested in investigating the presence of acetyl groups on all the Nterminal tail lysine residues of H3 and H4 in stationary and in exponential phase in S. cerevisiae, to elucidate the role that acetylation may play in transcriptional shutdown and compaction of the chromatin fiber during semi-quiescence
Summary
The involvement of histone acetylation in facilitating gene expression is well-established, in the case of histones H3 and H4 It was previously shown in Saccharomyces cerevisiae that gene expression was significantly down-regulated and chromatin more condensed in stationary phase compared to exponential phase. We were interested in establishing the acetylation state of histone H3 and H4 in stationary and in exponential phase, since the regulation of this modification could contribute to transcriptional shut-down and chromatin compaction during semi-quiescence. In 1963 Allfrey and colleagues showed that the acetylation of histone H3 and H4 alleviated the repressive effect of the histones on in vitro RNA synthesis in calf nuclei [1,2] This initial report saw the birth of epigenetics and an appreciation that covalent modifications of histones provided a mechanism whereby DNA function could be regulated. We were interested in investigating the presence of acetyl groups on all the Nterminal tail lysine residues of H3 and H4 in stationary and in exponential phase in S. cerevisiae, to elucidate the role that acetylation may play in transcriptional shutdown and compaction of the chromatin fiber during semi-quiescence
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