Defective bypass replication of a leading strand cyclobutane thymine dimer in xeroderma pigmentosum variant cell extracts.
Xeroderma pigmentosum variant (XP-V) is an inherited disorder resulting in hypersensitivity to the cytotoxic, mutagenic, and carcinogenic effects of UV light. There is evidence suggesting that XP-V cells carry a defect in the replication of UV-induced DNA damage, leading to mutations in genes, e.g., proto-oncogenes and tumor suppressor genes, of exposed skin cells. Using an in vitro assay to quantitatively evaluate replication of the most prevalent UV-derived DNA lesion, the cis,syn-thymine dimer (T x T), we have recently found that a T x T located on the leading strand can be bypassed by a bona fide human replication fork but can also induce fork uncoupling with selective synthesis of the undamaged lagging strand (D. Svoboda and J-M. Vos, Proc. Natl. Acad. Sci. USA, 92: 11975-11979, 1995). We now report the application and further refinement of this sensitive assay to the replication of a T x T-containing template by XP-V cell-free extracts. In comparison to normal controls, a 10-26-fold deficiency in the bypass replication of T x T was observed in XP-V cell extracts. In contrast, the disease extracts were as competent as controls for replication of the undamaged TT plasmid and for leading T x T-induced fork uncoupling. Besides mismatch repair and nucleotide excision repair, the bypass replication defect of XP-V may represent a novel category of hereditary mutator phenotypes affecting DNA damage processing.
Svoboda
D L
DL
Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, 27599-7295, USA.
Briley
L P
LP
Vos
J M
JM
D002474:Cell-Free System; D004249:DNA Damage; D004261:DNA Replication / Q000528:radiation effects; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D019281:Dimerization; D006367:HeLa Cells; D006801:Humans; D016296:Mutagenesis; D011740:Pyrimidine Dimers / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D013941:Thymine / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D013997:Time Factors; D014466:Ultraviolet Rays; D014983:Xeroderma Pigmentosum / Q000235:genetics* / Q000378:metabolism
D011740:Pyrimidine Dimers; C059776:endodeoxyribonuclease MfeI; D015252:Deoxyribonucleases, Type II Site-Specific; D013941:Thymine