Novel relationships among DNA methylation, histone modifications and gene expression in Ascobolus.

By studying Ascobolus strains methylated in various portions of the native met2 gene or of the hph transgene, we generalized our previous observation that methylation of the downstream portion of a gene promotes its stable silencing and triggers the production of truncated transcripts which rarely extend through the methylated region. In contrast, methylation of the promoter region does not promote efficient gene silencing. The chromatin state of met2 methylated strains was investigated after partial micrococcal nuclease (MNase) digestion. We show that MNase sensitive sites present along the unmethylated regions are no longer observed along the methylated ones. These chromatin changes are not resulting from the absence of transcription. They are associated, in both met2 and hph, with modifications of core histones corresponding, on the N terminus of histone H3, to an increase of dimethylation of lysine 9 and a decrease of dimethylation of lysine 4. Contrary to other organisms, these changes are independent of the transcriptional state of the genes, and furthermore, no decrease in acetylation of histone H4 is observed in silenced genes.