title: Human DNA methyltransferase gene-transformed yeasts display an inducible flocculation inhibited by 5-aza-2'-deoxycytidine.
abstract: Mammalian DNA methyltransferases (DNMTs) play an important role in establishing and maintaining the proper regulation of epigenetic information. However, it remains unclear whether mammalian DNMTs can be functionally expressed in yeasts, which probably lack endogenous DNMTs. We cotransformed the budding yeast Saccharomyces cerevisiae with the human DNMT1 gene, which encodes a methylation maintenance enzyme, and the DNMT3A/3B genes, which encode de novo methylation enzymes, in an expression vector also containing the GAL1 promoter, which is induced by galactose, and examined the effects of the DNMT inhibitor 5-aza-2'-deoxycytidine (5AZ) on cell growth. Transformed yeast strains grown in galactose- and glucose-containing media showed growth inhibition, and their growth rate was unaffected by 5AZ. Conversely, 5AZ, but not 2'-deoxycytidine, dose-dependently interfered with the flocculation exhibited by DNMT-gene transformants grown in glucose-containing medium. Further investigation of the properties of this flocculation indicated that it may be dependent on the expression of a Flocculin-encoding gene, FLO1. Taken together, these findings suggest that DNMT-gene transformed yeast strains functionally express these enzymes and represent a useful tool for in vivo screening for DNMT inhibitors.
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan. Electronic address: sugiyama@nihs.go.jp.
authors.forename: Kei-Ichi
authors.initials: K
authors.lastname: Sugiyama
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Makiko
authors.initials: M
authors.lastname: Takamune
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Furusawa
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Masamitsu
authors.initials: M
authors.lastname: Honma
mesh_terms: D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D003470:Culture Media / Q000378:metabolism; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000090205:DNA Methyltransferase 3A; D000077209:Decitabine; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D005427:Flocculation; D005686:Galactokinase / Q000235:genetics; D005947:Glucose / Q000378:metabolism; D006801:Humans; D037241:Mannose-Binding Lectins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D014170:Transformation, Genetic; D000097584:DNA Methyltransferase 3B
chemical_list: D001374:Azacitidine; D003470:Culture Media; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; D000090205:DNA Methyltransferase 3A; D000077209:Decitabine; D004791:Enzyme Inhibitors; D005686:Galactokinase; D005947:Glucose; D037241:Mannose-Binding Lectins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D000097584:DNA Methyltransferase 3B; C000717967:DNMT3A protein, human; C087578:FLO1 protein, S cerevisiae; C000619380:DNMT1 protein, human; C524219:GAL1 protein, S cerevisiae
keywords: 5-aza-2′-deoxycytidine (5AZ); DNA methyltransferase (DNMT); FLO1; Flocculation