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Dynamic Changes in Histone Modifications Precede de novo DNA Methylation in Oocytes

ABSTRACT: Erasure and subsequent re-instatement of DNA methylation in the germline, especially at imprinted CpG islands (CGIs), is crucial to embryogenesis in mammals. The mechanisms underlying DNA methylation establishment remain poorly understood, but a number of post-translational modifications of histones are implicated in antagonizing or recruiting the de novo DNA methylation complex. In mouse oogenesis, DNA methylation establishment occurs on a largely unmethylated genome and in non-dividing cells, making it a highly informative model for examining how histone modifications can shape the DNA methylome. Using a chromatin immunoprecipitation and genome-wide sequencing (ChIP-Seq) protocol optimized for low cell numbers and novel techniques for isolating primary and growing oocytes, profiles were generated for histone modifications implicated in promoting or inhibiting DNA methylation. CGIs destined for DNA methylation show reduced protective H3K4me2 and H3K4me3 in both primary and growing oocytes, while permissive H3K36me3 increases specifically at these CGIs in growing oocytes. Methylome profiling of oocytes deficient in H3K4 demethylases KDM1A or KDM1B indicated that removal of H3K4 methylation is necessary for proper methylation establishment at CGIs. This work represents the first systematic study performing ChIP-Seq in oocytes, and shows that histone remodeling in the mammalian oocyte helps direct de novo DNA methylation events.

ORGANISM(S): Mus musculus

PROVIDER: GSE74549 | GEO | 2015/12/02

SECONDARY ACCESSION(S): PRJNA300683

REPOSITORIES: GEO

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Project description:Dramatic changes in chromatin structure and histone modification occur during oocyte growth, as well as a global cessation of transcription. The role of histone modifications in these processes is poorly understood. We report the effect of conditionally deleting Hdac1 and Hdac2 on oocyte development. Deleting either gene has little or no effect on oocyte development, whereas deleting both genes results in follicle development arrest at the secondary follicle stage. This developmental arrest is accompanied by substantial perturbation of the transcriptome and a global reduction in transcription even though histone acetylation is markedly increased. There is no apparent change in histone repressive marks but there is a pronounced decrease in histone H3K4 methylation, an activating mark. The decrease in H3K4 methylation is likely due to increased expression of Kdm5b because RNAi-mediated targeting of Kdm5b in double mutant oocytes results in an increase in H3K4 methylation. An increase in TRP53 acetylation also occurs in mutant oocytes and may contribute to the observed increased incidence of apoptosis. Taken together, these results suggest seminal roles of acetylation of histone and non-histone proteins in oocyte development. We used microarrays to detail the global programme of gene expression underlying oocyte development and identified distinct classes of regulated genes during this process. Total RNA was extracted from 80 oocytes isolated from mice 12 days-of-age using the PicoPure RNA kit (Arcturus), amplified with the Ovation Pico WTA system (NuGen), and then fragmented and labeled with the FL-OvationTM cDNA Biotin Module V2 (NuGen). Four independent biological replicates were hybridized to Affymetrix GeneChip Mouse 1.1 ST microarrays (http://www.affymetrix.com/). Raw microarray data were analyzed as previously described using MAS5, GeneSpring v7, SAM and EASE software (56). A 1.4-fold cutoff was used for EASE analysis; four biological replicates provide sufficient statistical power and confidence to detect a 1.4-fold change in transcript abundance

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Dynamic Changes in Histone Modifications Precede de novo DNA Methylation in Oocytes

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