title: Molecular basis for 5-carboxycytosine recognition by RNA polymerase II elongation complex.
abstract: DNA methylation at selective cytosine residues (5-methylcytosine (5mC)) and their removal by TET-mediated DNA demethylation are critical for setting up pluripotent states in early embryonic development. TET enzymes successively convert 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC), with 5fC and 5caC subject to removal by thymine DNA glycosylase (TDG) in conjunction with base excision repair. Early reports indicate that 5fC and 5caC could be stably detected on enhancers, promoters and gene bodies, with distinct effects on gene expression, but the mechanisms have remained elusive. Here we determined the X-ray crystal structure of yeast elongating RNA polymerase II (Pol II) in complex with a DNA template containing oxidized 5mCs, revealing specific hydrogen bonds between the 5-carboxyl group of 5caC and the conserved epi-DNA recognition loop in the polymerase. This causes a positional shift for incoming nucleoside 5'-triphosphate (NTP), thus compromising nucleotide addition. To test the implication of this structural insight in vivo, we determined the global effect of increased 5fC/5caC levels on transcription, finding that such DNA modifications indeed retarded Pol II elongation on gene bodies. These results demonstrate the functional impact of oxidized 5mCs on gene expression and suggest a novel role for Pol II as a specific and direct epigenetic sensor during transcription elongation.
authors.affiliation: Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Lanfeng
authors.initials: L
authors.lastname: Wang
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Liang
authors.initials: L
authors.lastname: Xu
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Rui
authors.initials: R
authors.lastname: Xiao
authors.affiliation: Department of Chemistry, Department of Biochemistry and Molecular Biology, and Institute for Biophysical Dynamics, Howard Hughes Medical Institute, The University of Chicago, Chicago, Illinois 60637, USA.
authors.forename: Xingyu
authors.initials: X
authors.lastname: Lu
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Liang
authors.initials: L
authors.lastname: Chen
authors.affiliation: Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Jenny
authors.initials: J
authors.lastname: Chong
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Hairi
authors.initials: H
authors.lastname: Li
authors.affiliation: Department of Chemistry, Department of Biochemistry and Molecular Biology, and Institute for Biophysical Dynamics, Howard Hughes Medical Institute, The University of Chicago, Chicago, Illinois 60637, USA.
authors.forename: Chuan
authors.initials: C
authors.lastname: He
authors.affiliation: Department of Cellular and Molecular Medicine, School of Medicine, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Xiang-Dong
authors.initials: XD
authors.lastname: Fu
authors.affiliation: Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: Dong
authors.initials: D
authors.lastname: Wang
mesh_terms: D044503:5-Methylcytosine / Q000031:analogs & derivatives; D018360:Crystallography, X-Ray; D003596:Cytosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D019175:DNA Methylation; D004260:DNA Repair; D044127:Epigenesis, Genetic; D006860:Hydrogen Bonding; D007700:Kinetics; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D013698:Templates, Genetic; D044502:Thymine DNA Glycosylase / Q000378:metabolism; D061805:Transcription Elongation, Genetic
chemical_list: C560974:5-carboxylcytosine; C560973:5-formylcytosine; C011865:5-hydroxymethylcytosine; D044503:5-Methylcytosine; D003596:Cytosine; D012319:RNA Polymerase II; D044502:Thymine DNA Glycosylase