Mammalian translesion DNA synthesis across an acrolein-derived deoxyguanosine adduct. Participation of DNA polymerase eta in error-prone synthesis in human cells.
alpha-OH-PdG, an acrolein-derived deoxyguanosine adduct, inhibits DNA synthesis and miscodes significantly in human cells. To probe the cellular mechanism underlying the error-free and error-prone translesion DNA syntheses, in vitro primer extension experiments using purified DNA polymerases and site-specific alpha-OH-PdG were conducted. The results suggest the involvement of pol eta in the cellular error-prone translesion synthesis. Experiments with xeroderma pigmentosum variant cells, which lack pol eta, confirmed this hypothesis. The in vitro results also suggested the involvement of pol iota and/or REV1 in inserting correct dCMP opposite alpha-OH-PdG during error-free synthesis. However, none of translesion-specialized DNA polymerases catalyzed significant extension from a dC terminus when paired opposite alpha-OH-PdG. Thus, our results indicate the following. (i) Multiple DNA polymerases are involved in the bypass of alpha-OH-PdG in human cells. (ii) The accurate and inaccurate syntheses are catalyzed by different polymerases. (iii) A modification of the current eukaryotic bypass model is necessary to account for the accurate bypass synthesis in human cells.
Yang
In-Young
IY
Laboratory of Chemical Biology, Department of Pharmacological Sciences, State University of New York at Stony Brook, 11794-8651, USA.
Miller
Holly
H
Wang
Zhigang
Z
Frank
Ekaterina G
EG
Ohmori
Haruo
H
Hanaoka
Fumio
F
Moriya
Masaaki
M
D000171:Acrolein / Q000494:pharmacology*; D002460:Cell Line; D002461:Cell Line, Transformed; D004247:DNA / Q000096:biosynthesis* / Q000737:chemistry; D018736:DNA Adducts; D017931:DNA Primers / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008956:Models, Chemical; D015345:Oligonucleotide Probes / Q000494:pharmacology; D009841:Oligonucleotides / Q000494:pharmacology; D010957:Plasmids / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D000099270:Y-Family DNA Polymerases
D000171:Acrolein; D004247:DNA; D018736:DNA Adducts; D017931:DNA Primers; D004259:DNA-Directed DNA Polymerase; D003849:Deoxyguanosine; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D018809:Proliferating Cell Nuclear Antigen; C424742:OH-1,-N(2)-propano-2'-deoxyguanosine; C404302:POLK protein, human; D000099270:Y-Family DNA Polymerases; C112338:Rad30 protein