Binding of Histone H1 to DNA Is Indifferent to Methylation at CpG Sequences

Abstract

The possibility that histone H1 binds preferentially to DNA containing 5-methylcytosine in the dinucleotide CpG is appealing, as it could help to explain the repressive effects of methylation on gene activity. In this study, the affinity of purified H1 for methylated and non-methylated DNA sequences has been tested using both naked DNA and chromatin. Based on a variety of assays (bandshifts, filter-binding assays, Southwestern blots, and nuclease sensitivity assays), we conclude that H1 has no significant preference for binding to naked methylated DNA. Similarly, H1 showed the same affinities for methylated and non-methylated DNA when assembled into chromatin in a Xenopus oocyte extract. Thus potential cooperative interaction of H1 with polynucleosomal complexes is not enhanced by the presence of DNA methylation.The major DNA modification in vertebrates is methylation at the 5 position of cytosine in the dinucleotide CpG. About 60 -90% of genomic CpGs contain 5-methyl cytosine (m 5 C). Transfection assays with methylated genes have shown that CpG methylation often causes repression of transcription (1-4). The severity of repression depends on a number of parameters, particularly the location of methylation relative to the promoter (1, 5; but see Ref. 6), the local density of methyl-CpG pairs, and the strength of the promoter under test (7). In general, high density methylation strongly inhibits transcription, whereas low density methylation can only inhibit weak promoters.Several lines of evidence suggest that proteins that bind preferentially to methylated DNA are important components of the repression mechanism. Indeed, binding of the methyl-CpGbinding protein (MeCP) 1 MeCP1 (8) shows the same dependence on local density of methylation as does transcriptional repression (4, 7). Methylation-associated transcriptional repression may also arise by other routes. CpG methylation is known to prevent binding of some transcription factors, and this is likely to contribute to repression in some cases (9, 10). Another potential cause of methylation-mediated repression is direct alteration of chromatin structure due to the presence of m 5 C. Involvement of chromatin in mediating the effects of DNA methylation is suggested by the experiments of Buschhausen et al. (3,11). In addition, Keshet et al. found that methylated DNA is preferentially assembled into nuclease-resistant chromatin after transfection (12). While these results could be explained by interaction between known methyl-CpG-binding proteins and chromatin, it is also possible that an ubiquitous component of chromatin, for example histones, interacts differentially with methylated DNA leading to an altered higher order structure that is incompatible with gene expression.Attempts to detect an effect of DNA methylation on nucleosomal cores have not been successful (13,14). The linker histone H1, however, is an attractive candidate for mediator of the effects of methylation on chromatin. H1 plays a role in chromatin condensation (15, 16). It is depleted in CpG i...

Histone H1 Is Dispensable for Methylation-Associated Gene Silencing in Ascobolus immersusand Essential for Long Life Span

“…(iii) H1 is present in both methylated and nonmethylated chromatin, as indicated by the observation that similar increases in chromatin accessibility were observed in the different strains lacking H1, independently of the methylation state. This finding is consistent with observations made in vitro showing that H1 does not bind preferentially to methylated DNA (5,35).…”

“…Further in vitro studies led to contradictory results. While H1 was reported by McArthur and Thomas (31) to bind preferentially to methylated DNA, Campoy et al (5) and Nightingale and Wolffe (35) concluded that binding of H1 was indifferent to methylation in chromatin reconstitution experiments. It is difficult to make decisive conclusions from these in vitro studies, since factors playing an important role in the assembly of cellular chromatin may be missing in these assays.…”

Periodic DNA Methylation in Maize Nucleosomes and Demethylation by Environmental Stress

“…One such signal has been proposed to be m 5 C, but the experimental evidence is controversial. For example, opposite results as to preferential binding of histone H1 to methylated DNA have been reported (32,33), and currently the interaction between histones and methylated DNA is in general not considered to be specific. However, under certain circumstances, nucleosome assembly appears to depend on CpG methylation (31,34).…”

Transcriptional Augmentation: Modulation of Gene Expression by Scaffold/Matrix-Attached Regions (S/MAR Elements)

“…Shulman Which are the domain-opening events that are triggered by regional demethylation? The possibility that histone H1 binds preferentially to DNA containing 5-methylcytosine in the dinucleotide CpG and would cause domain opening after its release by demethylation (IVA2) is appealing but is without experimental support (Campoy et al, 1995). An alternative model is based on methylation-specific interactions with methyl CpG binding proteins (MeCPs) that are used to repress certain genes.…”