Abstract
Histone methylation is a prominent feature of eukaryotic chromatin that modulates multiple aspects of chromosome function. Methyl modification can occur on several different amino acid residues and in distinct mono-, di-, and tri-methyl states. However, the interplay among these distinct modification states is not well understood. Here we investigate the relationships between dimethyl and trimethyl modifications on lysine 9 of histone H3 (H3K9me2 and H3K9me3) in the adult Caenorhabditis elegans germ line. Simultaneous immunofluorescence reveals very different temporal/spatial localization patterns for H3K9me2 and H3K9me3. While H3K9me2 is enriched on unpaired sex chromosomes and undergoes dynamic changes as germ cells progress through meiotic prophase, we demonstrate here that H3K9me3 is not enriched on unpaired sex chromosomes and localizes to all chromosomes in all germ cells in adult hermaphrodites and until the primary spermatocyte stage in males. Moreover, high-copy transgene arrays carrying somatic-cell specific promoters are highly enriched for H3K9me3 (but not H3K9me2) and correlate with DAPI-faint chromatin domains. We further demonstrate that the H3K9me2 and H3K9me3 marks are acquired independently. MET-2, a member of the SETDB histone methyltransferase (HMTase) family, is required for all detectable germline H3K9me2 but is dispensable for H3K9me3 in adult germ cells. Conversely, we show that the HMTase MES-2, an E(z) homolog responsible for H3K27 methylation in adult germ cells, is required for much of the germline H3K9me3 but is dispensable for H3K9me2. Phenotypic analysis of met-2 mutants indicates that MET-2 is nonessential for fertility but inhibits ectopic germ cell proliferation and contributes to the fidelity of chromosome inheritance. Our demonstration of the differential localization and independent acquisition of H3K9me2 and H3K9me3 implies that the trimethyl modification of H3K9 is not built upon the dimethyl modification in this context. Further, these and other data support a model in which these two modifications function independently in adult C. elegans germ cells.
Highlights
Chromatin methylation is a complex and dynamic feature of eukaryotic chromosomes
In this work we investigate the relationships between the dimethyl and trimethyl modifications on lysine 9 of histone H3 (H3K9me2 and H3K9me3)
Despite the fact that H3K9me2 and H3K9me3 modify the same amino acid residue, our work reveals that these marks exhibit very different localization patterns in the adult C. elegans germ line and become concentrated on chromatin with different properties
Summary
Chromatin methylation is a complex and dynamic feature of eukaryotic chromosomes. Methylation can occur on numerous different sites on the histone H3 and H4 subunits. At some amino acid residues, such as lysine 4 and lysine 36 of histone H3, the presence of methyl modifications is correlated with active gene expression (reviewed in [1]). The diversity of methyl modifications is partially reflected in the large number of histone methyltransferases (HMTases) encoded by genomes. In addition to modifying a diverse range of histone residues, methyl marks occur in distinct mono-, di- and tri- methyl states. Histones bearing the di- and tri- methyl modifications at a given amino acid residue exhibit similar genomic distributions (e.g. H3K9 methylation in S. pombe [5]), or are dependent on the same HMTase (e.g. H3K27me and H3K27me dependence on MES-2 in C. elegans germ cells [6]). In many cases the relationships between the distinct methylation states and the HMTases that are responsible for generating them are poorly understood
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