A single (6-4) photoproduct inhibits plasmid DNA replication in xeroderma pigmentosum variant cell extracts.
The human skin cancer-prone disease xeroderma pigmentosum variant (XPV) results from a mutation in the human RAD30 gene, which encodes the lesion bypass DNA polymerase eta. XPV cells are characterized by delayed completion of DNA replication and increased mutagenesis following UV-irradiation. Using extracts of an XPV lymphoblast cell line (GM2449C) that has a truncating mutation in the RAD30 gene, we investigated the effect of a (6-4) photoproduct and a cyclobutane pyrimidine dimer (CPD), at a unique -TT- site on either the leading or lagging strand, on plasmid DNA replication. Compared to normal cell extracts, XPV cell extracts have a reduced capacity to carry out complete replication of DNA containing either a (6-4) photoproduct or a CPD on the leading strand, whereas there is little difference between the two cell extracts in replication of DNA containing a lesion on the lagging strand. Inhibition of replication in the presence of a (6-4) photoproduct is attributed to arrest of nascent DNA strand synthesis at the lesion site; in XPV cell extracts, the proportion of arrested products is increased compared to that of normal cell extracts. These results are consistent with a requirement for functional DNA polymerase eta in the replication of a double-stranded plasmid containing either a (6-4) photoproduct or a CPD, on the leading but not the lagging strand.
Yao
J
J
Department of Environmental Health, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
Dixon
K
K
Carty
M P
MP
D001483:Base Sequence; D002457:Cell Extracts; D004247:DNA / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D066298:In Vitro Techniques; D010957:Plasmids; D011740:Pyrimidine Dimers / Q000494:pharmacology*; D014983:Xeroderma Pigmentosum / Q000235:genetics*
D002457:Cell Extracts; D011740:Pyrimidine Dimers; C058406:pyrimidine-pyrimidone dimer; D004247:DNA; C020102:endodeoxyribonuclease DpnI; D015252:Deoxyribonucleases, Type II Site-Specific