title: Cell division is required for de novo methylation of CpG islands in bladder cancer cells.
abstract: Cell division is essential for tumor development and progression. Methylation-mediated silencing caused by aberrant de novo methylation of CpG islands located in the promoter regions of growth regulatory genes occurs frequently in human cancers. We investigated the relationship between cell division and de novo methylation to determine whether de novo methylation can occur in the absence of cell division in cancer cells. We treated T24 bladder carcinoma cells with 5-Aza-2'-deoxycytidine to induce a transient demethylation and then compared the timing and kinetics of remethylation of the p16 gene locus under conditions of either G(0)-G(1) growth arrest induced by serum starvation and confluence or continuous cell proliferation in complete medium. Variable levels of remethylation were detected in CpG poor regions of DNA, as well as repetitive DNA elements in the absence of cell division, yet no remethylation occurred at CpG islands under these conditions. This correlated with continuous expression of p16 protein in these cells. DNA methyltransferase (DNMT)1 and DNMT3b3 proteins were undetectable in 5-Aza-2'-deoxycytidine-treated and untreated nondividing cells, and their mRNA transcripts were down-regulated in these cells. Although DNMT3a mRNA levels were also reduced, they recovered to original levels in nondividing cells after drug treatment. Our results suggest that cell division is required for de novo methylation of CpG islands and that DNMT3a may play a role in methylating CpG poor regions or repetitive DNA elements outside of the S phase of the cell cycle.
authors.affiliation: Urologic Cancer Research Laboratory, Department of Biochemistry and Molecular Biology, University of Southern California/Norris Comprehensive Cancer Center, 1441 Eastlake Avenue, Los Angeles, CA 90089-9181, USA.
authors.forename: Mihaela
authors.initials: M
authors.lastname: Velicescu
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Weisenberger
authors.forename: Felicidad A
authors.initials: FA
authors.lastname: Gonzales
authors.forename: Yvonne C
authors.initials: YC
authors.lastname: Tsai
authors.forename: Carvell T
authors.initials: CT
authors.lastname: Nguyen
authors.forename: Peter A
authors.initials: PA
authors.lastname: Jones
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002295:Carcinoma, Transitional Cell / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002455:Cell Division / Q000502:physiology; D018899:CpG Islands / Q000187:drug effects / Q000502:physiology; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000096:biosynthesis / Q000235:genetics; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000096:biosynthesis / Q000235:genetics; D019175:DNA Methylation / Q000187:drug effects; D000090205:DNA Methyltransferase 3A; D000077209:Decitabine; D015536:Down-Regulation; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D019942:Genes, p16 / Q000502:physiology; D006801:Humans; D007700:Kinetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D000964:Antimetabolites, Antineoplastic; D019941:Cyclin-Dependent Kinase Inhibitor p16; C000717967:DNMT3A protein, human; D012333:RNA, Messenger; D000077209:Decitabine; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D001374:Azacitidine