Abstract

BackgroundDNA methylation, histone modifications and nucleosome occupancy act in concert for regulation of gene expression patterns in mammalian cells. Recently, G9a, a H3K9 methyltransferase, has been shown to play a role in establishment of DNA methylation at embryonic gene targets in ES cells through recruitment of de novo DNMT3A/3B enzymes. However, whether G9a plays a similar role in maintenance of DNA methylation in somatic cells is still unclear.ResultsHere we show that G9a is not essential for maintenance of DNA methylation in somatic cells. Knockdown of G9a has no measurable effect on DNA methylation levels at G9a-target loci. DNMT3A/3B remain stably anchored to nucleosomes containing methylated DNA even in the absence of G9a, ensuring faithful propagation of methylated states in cooperation with DNMT1 through somatic divisions. Moreover, G9a also associates with nucleosomes in a DNMT3A/3B and DNA methylation-independent manner. However, G9a knockdown synergizes with pharmacologic inhibition of DNMTs resulting in increased hypomethylation and inhibition of cell proliferation.ConclusionsTaken together, these data suggest that G9a is not involved in maintenance of DNA methylation in somatic cells but might play a role in re-initiation of de novo methylation after treatment with hypomethylating drugs, thus serving as a potential target for combinatorial treatments strategies involving DNMTs inhibitors.

Highlights

  • DNA methylation, histone modifications and nucleosome occupancy act in concert for regulation of gene expression patterns in mammalian cells

  • We show that G9a is not required for anchoring of DNMT3A/3B to nucleosomes in methylated chromatin regions and for maintenance of DNA methylation in somatic cells

  • G9a strongly associates with polynucleosomes similar to DNMT3A/3B To test the role of G9a, the euchromatic histone methyltransferase, in the strong nucleosome anchoring manifested by DNMT3A/3B, we examined their nucleosomal binding patterns using sucrose density gradient analysis

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Summary

Introduction

DNA methylation, histone modifications and nucleosome occupancy act in concert for regulation of gene expression patterns in mammalian cells. G9a, a H3K9 methyltransferase, has been shown to play a role in establishment of DNA methylation at embryonic gene targets in ES cells through recruitment of de novo DNMT3A/3B enzymes. DNA methylation is an essential epigenetic gene silencing mechanism which interplays with histone modifications and nucleosome occupancy for regulation of tissue-specific gene expression patterns and chromatin architecture in mammalian cells [1]. The de novo DNA methytransferases, DNMT3A/3B, primarily establish the methylation patterns during embryonic development [4] and later maintain them in differentiated tissues through cooperation with the maintenenace DNA methyltransferase, DNMT1 [5,6,7]. The mechanisms responsible for preferential targeting of DNMT3A/3B to such methylated regions are still poorly understood

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