NDLI: Rapid gene-specific repair of cisplatin lesions at the human DUG/DHFR locus comprising the divergent upstream gene and dihydrofolate reductase gene during early G1 phase of the cell cycle assayed by using the exonucleolytic activity of T4 DNA polymerase.

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Rapid gene-specific repair of cisplatin lesions at the human DUG/DHFR locus comprising the divergent upstream gene and dihydrofolate reductase gene during early G1 phase of the cell cycle assayed by using the exonucleolytic activity of T4 DNA polymerase.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Rampino, N. J. Bohr, V. A.
Description	Author Affiliation: Rampino NJ ( Laboratory of Molecular Genetics, National Institute on Aging, National Institutes of Health, Baltimore, MD 21224.);
Abstract	A novel assay to detect strand-specific DNA repair after cellular exposure to cisplatin at IC50 levels, is used to measure rapid repair in the divergent upstream gene (DUG), a human MutS homolog, and in the bidirectional promoter for dihydrofolate reductase gene (DHFR) and the contiguous upstream DUG. Single-stranded DNA capable of hybridizing to gene-specific probes is generated enzymatically by the 3'-5' exonuclease activity of T4 DNA polymerase. The presence of cisplatin lesions inhibit the exonucleolytic activity of T4 DNA polymerase and block the formation of single-stranded DNA. This decreases the amount of complementary sequence produced when assayed by gene-specific probe hybridization. With the progression of repair, increasing quantities of single-stranded DNA become available for probe hybridization. This assay was applied to human A2780 ovarian carcinoma cells treated with cisplatin at the beginning of G1 phase. A dose-response experiment showed that the assay was applicable down to cisplatin concentrations of 2.5 microM. To assay for strand-specific gene repair, the synchronized cells were treated with cisplatin and then allowed time to repair in drug-free medium. Extensive removal of cisplatin lesions after 2 hr of cellular repair during early G1 phase in the DUG and the DUG/DHFR promoter was measured, with no evidence of repair in the unexpressed delta-globin gene. The extent of preferential DNA repair was much more distinct than has been observed previously at high-drug dosage in asynchronous cells.
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	23
Volume Number	91
Language	English
Publisher	National Academy of Sciences
Publisher Date	1994-12-01
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Cisplatin Toxicity DNA Repair DNA-Directed DNA Polymerase Promoter Regions, Genetic Tetrahydrofolate Dehydrogenase Genetics Viral Proteins Cell Cycle Cells, Cultured DNA Damage Globins In Vitro Techniques Kinetics Restriction Mapping Multidisciplinary
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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