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)....

In the earliest stages of antigen receptor assembly, D and J segments of the Ig heavy chain and T cell receptor ␤ loci are recombined in B and T cells, respectively, whereas the V segments are not. Distinct distribution patterns of various histone modifications and the nucleosome-remodeling factor BRG1 are found at ''active'' (DJ) and ''inactive'' (V) regions. Striking ''hotspots'' of histone H3 dimethylated at lysine 4 (di-Me H3-K4) are localized at the ends of the active DJ domains of both the Ig heavy chain and T cell receptor ␤ loci. BRG1 is not localized to specific sequences, as it is with transcriptional initiation, but rather associates with the entire active locus in a pattern that mirrors acetylation of histone H3. Within some inactive loci marked by H3-K9 dimethylation, two distinct levels of methylation are found in a nonrandom genesegment-specific pattern. We suggest that the hotspots of di-Me H3-K4 are important marks for locus accessibility. The specific patterns of modification imply that the regulation of V(D)J recombination involves recruitment of specific methyltransferases in a localized manner.T he chromatin in the nucleus of eukaryotic cells is regulated to permit or exclude access of the enzymatic machinery for processes such as transcription and recombination. Specific regulatory sequences in the DNA are ultimately responsible for this regulation, serving as binding sites for proteins or protein complexes that recruit specific chromatin-modifying activities. In the case of transcriptional regulation, specific DNA-binding activators or repressors recruit histone-modifying enzymes and nucleosome remodeling complexes, generating localized modifications of the chromatin that govern the access of the transcription machinery (reviewed in refs. 1 and 2). In addition to such localized chromatin modifications, there are developmentally regulated large-scale reorganizations of chromatin structure into active and inactive domains [e.g., ␤-globin genes in chickens (3) and the mating-type locus in Schizosaccharomyces pombe (4)] (reviewed in ref. 5).A variety of covalent histone modifications are associated with active or inactive chromatin (6-8). Acetylation of histones H3 and H4 is well known to mark transcriptionally active chromatin. Dimethylation of histone H3 on lysine 4 (di-Me H3-K4) is often observed at active loci in yeast (4), Tetrahymena (9), and chicken (3), although associations with silent loci have also been reported (10). In contrast, dimethylation of histone H3 on lysine 9 (di-Me H3-K9) is often, if not always, correlated with regions of silent inactive chromatin (11).Coordination of the series of DNA rearrangement events required to assemble Ig and TCR genes from component V, D, and J segments presents a highly complex regulatory problem (reviewed in refs. 12 and 13). There are seven structurally unique antigen receptor loci, some spanning 3 or 4 megabases, each composed of multiple V, J, and sometimes D segments, along with nonrearranging constant (C) gene segments. Rearrangement is ...

The ordered assembly of immunoglobulin and TCR genes by V(D)J recombination depends on the regulated accessibility of individual loci. We show here that the histone tails and intrinsic nucleosome structure pose significant impediments to V(D)J cleavage. However, alterations to nucleosome structure via histone acetylation or by stable hSWI/SNF-dependent remodeling greatly increase the accessibility of nucleosomal DNA to V(D)J cleavage. Moreover, acetylation and hSWI/SNF remodeling can act in concert on an individual nucleosome to achieve levels of V(D)J cleavage approaching those observed on naked DNA. These results are consistent with a model in which regulated recruitment of chromatin modifying activities is involved in mediating the lineage and stage-specific control of V(D)J recombination.