The pluripotency-determining gene Oct3/4 (also called Pou5f1) undergoes postimplantation silencing in a process mediated by the histone methyltransferase G9a. Microarray analysis now shows that this enzyme may operate as a master regulator that inactivates numerous early-embryonic genes by bringing about heterochromatinization of methylated histone H3K9 and de novo DNA methylation. Genetic studies in differentiating embryonic stem cells demonstrate that a point mutation in the G9a SET domain prevents heterochromatinization but still allows de novo methylation, whereas biochemical and functional studies indicate that G9a itself is capable of bringing about de novo methylation through its ankyrin domain, by recruiting Dnmt3a and Dnmt3b independently of its histone methyltransferase activity. These modifications seem to be programmed for carrying out two separate biological functions: histone methylation blocks target-gene reactivation in the absence of transcriptional repressors, whereas DNA methylation prevents reprogramming to the undifferentiated state.
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We are grateful to S. Tajima (Osaka University) for Dnmt3a and 3b expression vectors, K. Wright (University of South Florida) for the hemagglutinin-G9a expression vector and S. Pradhan (New England Biolabs) for pGEX-G9a. This work was supported by grants from the Israel Academy of Science (Y.B. and H.C.), Philip Morris USA Inc. and Philip Morris International (S.E.-L., Y.B. and H.C.), the National Institutes of Health (Y.B. and H.C.), the Israel Cancer Research Fund (Y.B. and H.C.) and the Prostate Cancer Foundation (H.C.).