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Analysis of the role of the histone methyltransferase G9a in transcriptional repression of introduced and endogenous retroelements in murine embryonic stem cells

University of British Columbia

DNA methylation and posttranslational histone modification are epigenetic marks that influence transcription of associated DNA. DNA methylation involves the direct addition of methyl groups to cytosine bases in DNA, and is associated with transcriptional repression. Histone modification can be associated with either activation or repression, and involves covalent addition of a wide variety of chemical groups to the N-terminal tails of the core histones. A relationship between DNA methylation of repetitive elements and methylation of H3 lysine 9 (H3K9) exists in Arabidopsis thaliana, where mutants in the histone methyltransferase (HMTase) KRYPTONITE (KYP) show a defect in CpNpG DNA methylation of retrotransposons. Similarly, in Neurospora crassa the H3K9 HMTase defective in methylation 5 (DIM-5) is required for DNA methylation of repetitive elements. In mice, the major H3K9 methyltransferases (MTase) in euchromatin are G9a and GLP; G9a has been shown to influence DNA methylation of a limited number of single copy genes, however, its effect on interspersed elements, such as endogenous retroviruses (ERVs) or exogenous retroviruses (XRVs) has not been characterized. Here, I show that G9a is responsible for H3K9 dimethylation (H3K9me2) and Kap-1 recruitment at ERVs, as well as H3K9me2 and H3K9me3 at XRVs. Moreover, G9a acted upstream of DNA methylation of both XRVs and ERVs. Mutation of G9a resulted in heightened expression of XRVs, but did not influence expression of ERVs. Analysis of the mechanism behind this histone modification-DNA methylation link revealed that recruitment of the DNA methyltransferase (DNMT) De novo methyltransferase 3a (Dnmt3a) was reduced at ERVs in the absence of G9a. Surprisingly, this recruitment was partially rescued by catalytically null G9a. Based on these observations, I conclude that G9a is targeted to ERVs and XRVs and marks these elements with H3K9me2. In addition, G9a acts upstream of DNA methylation and H3K9me3 at XRVs. The DNA methylation deficiency may be partly explained by recruitment of Dnmt3a independent of G9a’s catalytic activity. The differences observed between G9a activity at exogenous and ERVs suggest that developmental and/or evolutionary mechanisms place additional constraints on ERVs compared to XRVs.

Text

2010-01-21

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/18820

Medical Genetics

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/