DNA adducts of cis-diamminedichloroplatinum(II) and its trans isomer inhibit RNA polymerase II differentially in vivo.
The effects of cis-diamminedichloroplatinum(II) (cis-DDP) and trans-DDP adducts on mammalian transcription in vivo have been investigated. A plasmid containing the beta-galactosidase (beta-gal) reporter gene was modified with either of the two platinum compounds and transfected into human or hamster cell lines. A 2-3 fold higher level of transcription was observed in both cell lines from plasmids containing trans-DDP adducts as compared to plasmids modified by cis-DDP. This difference in transcriptional activity was not decreased in human and rodent nucleotide excision repair deficient cell lines, indicating that more efficient excision repair of the trans-DDP adducts was not the cause of its lower ability to block transcription in this assay. For this conclusion to be valid, it is assumed that trans-DDP adducts are repaired primarily by the nucleotide excision repair pathway, as is the case with the adducts of cis-DDP. The possibility that trans-DDP adducts are preferentially bypassed by RNA polymerase was examined by monitoring the elongation of beta-gal mRNA on damaged templates in vivo. Nascent beta-gal mRNA transcripts were recovered from excision repair deficient xeroderma pigmentosum A cells transfected with platinated plasmids, and the extent of RNA synthesis was measured by using ribonuclease protection. Fourfold more trans-DDP than cis-DDP adducts were required to inhibit transcription elongation by 63%. RNA polymerase II bypassed cis- and trans-DDP DNA adducts with efficiencies of 0-16% and 60-70%, respectively. These data provide insight into the differential toxicity of the two platinum isomers.
Mello
J A
JA
Department of Chemistry, Whitaker College of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge 02139, USA.
Lippard
S J
SJ
Essigmann
J M
JM
D000818:Animals; D016375:Antisense Elements (Genetics); D016466:CHO Cells; D002945:Cisplatin / Q000378:metabolism / Q000494:pharmacology*; D006224:Cricetinae; D004247:DNA / Q000187:drug effects / Q000378:metabolism*; D018736:DNA Adducts / Q000494:pharmacology*; D004249:DNA Damage; D015342:DNA Probes; D004260:DNA Repair; D015870:Gene Expression; D017930:Genes, Reporter / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D012319:RNA Polymerase II / Q000037:antagonists & inhibitors*; D016372:RNA, Antisense / Q000235:genetics; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
D016375:Antisense Elements (Genetics); D018736:DNA Adducts; D015342:DNA Probes; D016372:RNA, Antisense; C057270:cisplatin-DNA adduct; C034697:transplatin; D004247:DNA; D012319:RNA Polymerase II; D002945:Cisplatin