title: Transcriptional inhibition and mutagenesis induced by N-nitroso compound-derived carboxymethylated thymidine adducts in DNA.
abstract: N-nitroso compounds represent a common type of environmental and endogenous DNA-damaging agents. After metabolic activation, many N-nitroso compounds are converted into a diazoacetate intermediate that can react with nucleobases to give carboxymethylated DNA adducts such as N3-carboxymethylthymidine (N3-CMdT) and O(4)-carboxymethylthymidine (O(4)-CMdT). In this study, we constructed non-replicative plasmids carrying a single N3-CMdT or O(4)-CMdT, site-specifically positioned in the transcribed strand, to investigate how these lesions compromise the flow of genetic information during transcription. Our results revealed that both N3-CMdT and O(4)-CMdT substantially inhibited DNA transcription mediated by T7 RNA polymerase or human RNA polymerase II in vitro and in human cells. In addition, we found that N3-CMdT and O(4)-CMdT were miscoding lesions and predominantly directed the misinsertion of uridine and guanosine, respectively. Our results also suggested that these carboxymethylated thymidine lesions may constitute efficient substrates for transcription-coupled nucleotide excision repair in human cells. These findings provided important new insights into the biological consequences of the carboxymethylated DNA lesions in living cells.
authors.affiliation: Department of Chemistry, University of California, Riverside, CA 92521-0403, USA.
authors.forename: Changjun
authors.initials: C
authors.lastname: You
authors.affiliation: Department of Chemistry, University of California, Riverside, CA 92521-0403, USA.
authors.forename: Jianshuang
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Chemistry, University of California, Riverside, CA 92521-0403, USA.
authors.forename: Xiaoxia
authors.initials: X
authors.lastname: Dai
authors.affiliation: Department of Chemistry, University of California, Riverside, CA 92521-0403, USA Yinsheng.Wang@ucr.edu.
authors.forename: Yinsheng
authors.initials: Y
authors.lastname: Wang
mesh_terms: D002460:Cell Line; D018736:DNA Adducts / Q000737:chemistry*; D012321:DNA-Directed RNA Polymerases / Q000037:antagonists & inhibitors; D006801:Humans; D016296:Mutagenesis; D013936:Thymidine / Q000031:analogs & derivatives* / Q000737:chemistry; D014158:Transcription, Genetic
chemical_list: D018736:DNA Adducts; C000597427:N3-carboxymethylthymidine; C000597428:O4-carboxymethylthymidine; D012321:DNA-Directed RNA Polymerases; D013936:Thymidine