UV lesions located on the leading strand inhibit DNA replication but do not inhibit SV40 T-antigen helicase activity

DNA replication in eucaryotic cells involves a variety of proteins which synthesize the leading and lagging strands in an asymmetric coordinated manner. To analyse the effect of this asymmetry on the translesion synthesis of UV-induced lesions, we have incubated SV40 origin-containing plasmids with...

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Bibliographic Details
Published in:Mutation research. DNA repair 2000-02, Vol.459 (1), p.19-28
Main Authors: Veaute, Xavier, Mari-Giglia, Giuseppina, Lawrence, Christopher W, Sarasin, Alain
Format: Article
Language:English
Subjects:
Biological and medical sciences
Cyclobutane pyrimidine dimers
DNA helicase
Fundamental and applied biological sciences. Psychology
Helicase assay
In vitro replication
Molecular and cellular biology
Molecular genetics
Mutagenesis. Repair
Pyrimidine (6-4) pyrimidones
Simian virus 40
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Summary:DNA replication in eucaryotic cells involves a variety of proteins which synthesize the leading and lagging strands in an asymmetric coordinated manner. To analyse the effect of this asymmetry on the translesion synthesis of UV-induced lesions, we have incubated SV40 origin-containing plasmids with a unique site-specific cis,syn-cyclobutane dimer or a pyrimidine-pyrimidone (6-4) photoproduct on either the leading or lagging strand template with DNA replication-competent extracts made from human HeLa cells. Two dimensional agarose gel electrophoresis analyses revealed a strong blockage of fork progression only when the UV lesion is located on the leading strand template. Because DNA helicases are responsible for unwinding duplex DNA ahead of the fork and are then the first component to encounter any potential lesion, we tested the effect of these single photoproducts on the unwinding activity of the SV40 T antigen, the major helicase in our in vitro replication assay. We showed that the activity of the SV40 T-antigen helicase is not inhibited by UV-induced DNA lesions in double-stranded DNA substrate.
ISSN:0921-8777
1386-1476
DOI:10.1016/S0921-8777(99)00052-X