Effects of Tet-mediated oxidation products of 5-methylcytosine on DNA transcription in vitro and in mammalian cells.
5-methylcytosine (5-mC) is a well-characterized epigenetic regulator in mammals. Recent studies showed that Ten-eleven translocation (Tet) proteins can catalyze the stepwise oxidation of 5-mC to produce 5-hydroxymethylcytosine (5-HmC), 5-formylcytosine (5-FoC) and 5-carboxylcytosine (5-CaC). The exciting discovery of these novel cytosine modifications has stimulated substantial research interests about their roles in epigenetic regulation. Here we systematically examined the effects of the oxidized 5-mC derivatives on the efficiency and fidelity of DNA transcription using a recently developed competitive transcription and adduct bypass assay. Our results showed that, when located on the transcribed strand, 5-FoC and 5-CaC exhibited marginal mutagenic and modest inhibitory effects on DNA transcription mediated by single-subunit T7 RNA polymerase or multi-subunit human RNA polymerase II in vitro and in human cells. 5-HmC displayed relatively milder blocking effects on transcription, and no mutant transcript could be detectable for 5-HmC in vitro or in cells. The lack of considerable mutagenic effects of the oxidized 5-mC derivatives on transcription was in agreement with their functions in epigenetic regulation. The modest blocking effects on transcription suggested that 5-FoC and 5-CaC may function in transcriptional regulation. These findings provided new evidence for the potential functional interplay between cytosine methylation status and transcription.
Department of Chemistry, University of California, Riverside, California 92521-0403.
Changjun
C
You
Department of Chemistry, University of California, Riverside, California 92521-0403.
Debin
D
Ji
Department of Chemistry, University of California, Riverside, California 92521-0403.
Xiaoxia
X
Dai
Department of Chemistry, University of California, Riverside, California 92521-0403.
Yinsheng
Y
Wang
D044503:5-Methylcytosine / Q000096:biosynthesis*; D000818:Animals; D002460:Cell Line; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D010084:Oxidation-Reduction; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D044503:5-Methylcytosine