Braunstein M, Sobel R E, Allis C D, Turner B M, Broach J R
Department of Molecular Biology, Princeton University, New Jersey 08544, USA.
Mol Cell Biol. 1996 Aug;16(8):4349-56. doi: 10.1128/MCB.16.8.4349.
Heterochromatin in metazoans induces transcriptional silencing, as exemplified by position effect variegation in Drosophila melanogaster and X-chromosome inactivation in mammals. Heterochromatic DNA is packaged in nucleosomes that are distinct in their acetylation pattern from those present in euchromatin, although the role these differences play in the structure of heterochromatin or in the effects of heterochromatin on transcriptional activity is unclear. Here we report that, as observed in the facultative heterochromatin of the inactive X chromosome in female mammalian cells, histones H3 and H4 in chromatin spanning the transcriptionally silenced mating-type cassettes of the yeast Saccharomyces cerevisiae are hypoacetylated relative to histones H3 and H4 of transcriptionally active regions of the genome. By immunoprecipitation of chromatin fragments with antibodies specific for H4 acetylated at particular lysine residues, we found that only three of the four lysine residues in the amino-terminal domain of histone H4 spanning the silent cassettes are hypoacetylated. Lysine 12 shows significant acetylation levels. This is identical to the pattern of histone H4 acetylation observed in centric heterochromatin of D. melanogaster. These two observations provide additional evidence that the silent cassettes are encompassed in the yeast equivalent of metazoan heterochromatin. Further, mutational analysis of the amino-terminal domain of histone H4 in S. cerevisiae demonstrated that this observed pattern of histone H4 acetylation is required for transcriptional silencing. This result, in conjunction with prior mutational analyses of yeast histones H3 and H4, indicates that the particular pattern of nucleosome acetylation found in heterochromatin is required for its effects on transcription and is not simply a side effect of heterochromatin formation.
后生动物中的异染色质会诱导转录沉默，果蝇中的位置效应斑驳以及哺乳动物中的X染色体失活就是例证。异染色质DNA被包装在核小体中，其乙酰化模式与常染色质中的核小体不同，尽管这些差异在异染色质结构或异染色质对转录活性的影响中所起的作用尚不清楚。我们在此报告，正如在雌性哺乳动物细胞中失活的X染色体的兼性异染色质中所观察到的那样，酿酒酵母转录沉默的交配型基因座的染色质中的组蛋白H3和H4相对于基因组转录活性区域的组蛋白H3和H4发生了低乙酰化。通过用针对特定赖氨酸残基乙酰化的H4的抗体对染色质片段进行免疫沉淀，我们发现跨越沉默基因座的组蛋白H4氨基末端结构域的四个赖氨酸残基中只有三个发生了低乙酰化。赖氨酸12显示出显著的乙酰化水平。这与在黑腹果蝇着丝粒异染色质中观察到的组蛋白H4乙酰化模式相同。这两个观察结果提供了额外的证据，表明沉默基因座包含在酵母中相当于后生动物异染色质的结构中。此外，对酿酒酵母中组蛋白H4氨基末端结构域的突变分析表明，这种观察到的组蛋白H4乙酰化模式是转录沉默所必需的。这一结果与先前对酵母组蛋白H3和H4的突变分析相结合，表明在异染色质中发现的核小体乙酰化的特定模式是其对转录产生影响所必需的，而不仅仅是异染色质形成的副作用。
