Epigenetic regulation of X-linked cancer/germline antigen genes by DNMT1 and DNMT3b.
We examined the function of two key DNA methyltransferase (DNMT) enzymes in epigenetic regulation of X-linked cancer/germline (CG-X) antigen genes in human cancer cells, using MAGE-A1, NY-ESO-1, and XAGE-1 as models. In HCT116 cells, genetic knockout of DNMT1 caused moderate activation of CG-X genes, DNMT3b knockout had a negligible effect, and double knockout of both enzymes caused robust gene induction. Similarly, dual DNMT knockout caused dramatic hypomethylation of the MAGE-A1 and NY-ESO-1 promoters, DNMT1 knockout showed moderate hypomethylation, and DNMT3b knockout elicited only slight methylation changes. In contrast, both single and double knockout cells showed significant hypomethylation of the XAGE-1 promoter. RNA interference (RNAi) targeting of DNMT1 in HCT116 cells validated the results seen using genetic knockout cells; however, RNAi targeting of DNMT1 in a different colorectal cancer cell line revealed a greater independent role for DNMT1 in mediating CG-X gene repression and promoter methylation in other cell types. Notably, the histone H3 modification pattern at CG-X promoters was altered following DNMT knockout. DNMT1 or DNMT3b knockout reduced dimethylated lysine-9 (diMe-H3K9) levels, but did not significantly affect dimethylated lysine-4 (diMe-H3K4) or acetylated lysine-9 (Ac-H3-K9) levels. In contrast, dual DNMT1/3b knockout reduced the level of diMe-H3K9 and dramatically increased the levels of diMe-H3K4 and Ac-H3K9 at CG-X gene loci. In summary, DNMT1 and DNMT3b were found to perform both redundant and independent functions in epigenetic regulation of CG-X antigen genes in human cancer cells.
James
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Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Cancer Drug Center, Buffalo, New York, NY 14263, USA.
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Karpf
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D000951:Antigens, Neoplasm / Q000235:genetics*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D017931:DNA Primers; D044127:Epigenesis, Genetic; D015870:Gene Expression; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D050172:Genes, X-Linked / Q000235:genetics*; D006657:Histones; D006801:Humans; D058950:Melanoma-Specific Antigens; D008565:Membrane Proteins / Q000235:genetics; D009363:Neoplasm Proteins / Q000235:genetics; D011401:Promoter Regions, Genetic; D034741:RNA, Small Interfering; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation; D000097584:DNA Methyltransferase 3B
D000951:Antigens, Neoplasm; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D017931:DNA Primers; D006657:Histones; D058950:Melanoma-Specific Antigens; D008565:Membrane Proteins; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D000097584:DNA Methyltransferase 3B; C104948:CTAG1B protein, human; C413656:XAGE1A protein, human; C000619380:DNMT1 protein, human