Effects of endogenous DNA base lesions on transcription elongation by mammalian RNA polymerase II. Implications for transcription-coupled DNA repair and transcriptional mutagenesis.
The blockage of transcription elongation by RNA polymerase II (pol II) is thought to be a trigger for transcription-coupled repair in the pathway of nucleotide excision repair. Purified pol II and oligo(dC)-tailed templates containing a single non-bulky DNA lesion on the transcribed strand such as an apurinic/apyrimidinic (AP) site, uracil, or 8-oxoguanine (8-oxoG) were used for transcription elongation assays. In this system pol II could bypass both the AP site and uracil without pausing and insert cytosine opposite the AP site and either guanine or adenine opposite to uracil. Thus, the AP site on the DNA templates could lead to correct transcription only if depurination at guanine occurred, whereas uracil generated either the correct transcriptional product or an incorrect one with a G:C to A:T transition. In the case of 8-oxoG, pol II stalled at the lesion, but sometimes bypassed it and inserted a cytosine residue or the incorrect adenine residue leading to a G:C to T:A transversion. These findings indicate that 8-oxoG lesions caused a blockage of transcription elongation and/or the misincorporation of a ribonucleotide by pol II, implying the initiation of transcription-coupled repair of 8-oxoG and/or transcriptional mutagenesis.
Kuraoka
Isao
I
Laboratories for Organismal Biosystems, Graduate School of Frontier Biosciences, Osaka University and Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Corporation, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
Endou
Masaki
M
Yamaguchi
Yuki
Y
Wada
Tadashi
T
Handa
Hiroshi
H
Tanaka
Kiyoji
K
D000225:Adenine / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D003570:Cytidine Triphosphate / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D004260:DNA Repair; D004270:DNA, Circular; D006147:Guanine / Q000737:chemistry; D006151:Guanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008957:Models, Genetic; D016296:Mutagenesis; D012313:RNA / Q000378:metabolism; D012319:RNA Polymerase II / Q000737:chemistry* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014158:Transcription, Genetic
D004270:DNA, Circular; D006151:Guanosine; C046215:8-hydroxyguanosine; D006147:Guanine; D012313:RNA; D003570:Cytidine Triphosphate; D000255:Adenosine Triphosphate; D004247:DNA; D012319:RNA Polymerase II; D000225:Adenine