Identification of three histone methyltransferases in Drosophila: dG9a is a suppressor of PEV and is required for gene silencing.

Organization of chromatin structure and regulation of gene transcription are contingent on histone tail modifications. Regions of the genome packaged with nucleosomes that contain methyl histone H3 at lysine 9 (Me K9H3) strongly correlate with regions that are silenced for transcription. To date Su(var)3-9 is the only K9H3 specific enzyme characterized in Drosophila melanogaster. In this study, we describe the identification of three additional Drosophila genes that potentially encode K9H3 specific methyltransferases (HMTase) with homology to known mammalian proteins. By several criteria, including sequence alignments, phylogenic analyses, and enzyme activity of the protein, one of these is a homologue of the human G9a and hence, we name it dG9a. dG9a catalyzes the transfer of methyl groups to full-length histone H3 and to N-terminal H3 peptides that contain lysine 9, suggesting that the major target for dG9a is K9H3. Chromatin extracts prepared from a P-element insert mutation in dG9a display an altered K9H3 methylation profile. In addition, the dG9a mutant is a dominant suppressor of position-effect variegation (PEV), a heterochromatin-associated gene silencing phenomenon. Su(var)3-9 also suppresses PEV. The combined Su(var)3-9 and dG9a mutations have severe developmental defects suggesting an overlapping role for dG9a and Su(var)3-9 in the packaging of heterochromatin and gene silencing via a K9H3 methylation pathway.

A role for the Drosophila SU(VAR)3-9 protein in chromatin organization at the histone gene cluster and in suppression of position-effect variegation.

Mutations in the gene for Su(var)3-9 are dominant suppressors of position-effect variegation (PEV). We show that SU(VAR)3-9 is a chromatin-associated protein and identify the large multicopy histone gene cluster (HIS-C) as one of its target loci. The organization of nucleosomes over the entire HIS-C region is altered in Su(var)3-9 mutants and there is a concomitant increase in expression of the histone genes. SU(VAR)3-9 is a histone H3 methyltransferase and, using chromatin immunoprecipitation, we show that SU(VAR)3-9 is present at the HIS-C locus and that the histone H3 at the HIS-C locus is methylated. We propose that SU(VAR)3-9 is involved in packaging HIS-C into a distinct chromatin domain that has some of the characteristics of beta-heterochromatin. We suggest that methylation of histone H3 is important for the chromatin structure at HIS-C. The chromosomal deficiency for the HIS-C is also a suppressor of PEV. In contrast to what might be expected, we show that hemizygosity for the HIS-C locus leads to a substantial increase in the histone transcripts.