Lysine-79 of histone H3 is hypomethylated at silenced loci in yeast and mammalian cells: A potential mechanism for position-effect variegation

Abstract

Methylation of lysine-79 (K79) within the globular domain of histone H3 by Dot1 methylase is important for transcriptional silencing and for association of the Sir silencing proteins in yeast. Here, we show that the level of H3-K79 methylation is low at all Sir-dependent silenced loci but not at other transcriptionally repressed regions. Hypomethylation of H3-K79 at the telomeric and silent mating-type loci, but not the ribosomal DNA, requires the Sir proteins. Overexpression of Sir3 concomitantly extends the domain of Sir protein association and H3-K79 hypomethylation at telomeres. In mammalian cells, H3-K79 methylation is found at loci that are active for V(D)J recombination, but not at recombinationally inactive loci that are heterochromatic. These results suggest that H3-K79 methylation is an evolutionarily conserved marker of active chromatin regions, and that silencing proteins block the ability of Dot1 to methylate histone H3. Further, they suggest that Sir proteins preferentially bind chromatin with hypomethylated H3-K79 and then block H3-K79 methylation. This positive feedback loop, and the reverse loop in which H3-K79 methylation weakens Sir protein association and leads to further methylation, suggests a model for position-effect variegation. In eukaryotic cells, DNA is packaged along with histones and other nuclear proteins to form chromatin. Cytologically, chromatin can be broadly classified into condensed heterochromatin and decondensed euchromatin (1-4). In general, heterochromatin is associated with repetitive elements at telomeric and pericentric chromosomal regions, contains few genes, and replicates late in S phase. However, certain chromosomal regions can be either heterochromatic or euchromatic, depending on developmental or environmental conditions. Heterochromatin in divergent animal species (3, 4) and the fission yeast Schizosaccharomyces pombe (5) has a characteristic pattern of histone modifications, in which the N-terminal histone tails are virtually nonacetylated and lysine-9 (K9) of histone H3 is methylated. Specific nonhistone proteins (e.g., HP1) associate with nucleosomes in which histone H3 is methylated at lysine-9, thereby providing a physical difference between heterochromatin and euchromatin. In the budding yeast Saccharomyces cerevisiae, telomeric, silent mating-type (HM), and ribosomal DNA (rDNA) loci are considered to be heterochromatic (2, 3). These heterochromatic loci have histones that are essentially nonacetylated (6, 7) but are atypical in that histone H3-K9 is not methylated and proteins such as HP1 do not exist.In general, DNA in heterochromatin is relatively inaccessible to enzymatic probes and is inert for transcription and recombination, whereas euchromatic DNA is accessible and active for these processes. In Drosophila, translocation of euchromatic genes or integration of reporter genes next to heterochromatin results in variegated expression, a phenomenon in which genetically identical cells stochastically express or repress the gene in a heritable manner (1)....

Histone sumoylation is a negative regulator in Saccharomyces cerevisiae and shows dynamic interplay with positive-acting histone modifications

“…4A). Strikingly, this is the reverse correlation as previously observed for both H2B-associated ubiquitylation and H3 methylation (Ng et al 2003;Emre et al 2005). Further, we found that telomeric silencing of URA3 cassette expression (Aparicio et al 1991) is dramatically lowered in the ubc9ts strain (data not shown), which may indicate a role of histone sumoylation in promoting telomeric silencing.…”

“…Telomeric silencing in S. cerevisiae has long been associated with reduced levels of positive-acting histone modifications. This includes low acetylation, maintained through the action of the histone deacetylase Sir2 (Grunstein 1997;Rusche et al 2003), and low ubiquitylation and methylation (Bryk et al 2002;Ng et al 2003), maintained via the histone deubiquitylase Ubp10, which also serves to keep methylation levels low (Emre et al 2005). We detect slightly higher H2B-associated sumoylation at telomeres than at more internal chromosomal sites or at genes (Fig.…”

The histone methylase Set2p and the histone deacetylase Rpd3p repress meiotic recombination at the HIS4 meiotic recombination hotspot in Saccharomyces cerevisiae

“…We also examined the effects of eliminating Dot1p, an H3K79 histone methyltransferase (38,39). H3K79 methylation is at low levels at yeast telomeres (40), but occurs at high levels at activelytranscribed genes (41). As described below, we found that deletion of SET2 or RPD3 strongly stimulated HIS4 hotspot activity, whereas deletion of HDA1 had a more subtle stimulatory effect.…”

Regional centromeres in the yeast Candida lusitaniae lack pericentromeric heterochromatin

“…H3K79 methylation is associated with euchromatin and transcriptional activity (50)(51)(52), but paradoxically loss of H3K79 methylation perturbs heterochromatin formation in budding yeast and mouse cells (51,53). Symmetric methylation of H3R2 is also associated with euchromatin (54).…”