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Related Experiment Videos

An alternative eukaryotic DNA excision repair pathway

G A Freyer1, S Davey, J V Ferrer

  • 1Center for Radiological Research, Columbia University College of Physicians and Surgeons, New York, New York 10032, USA.

Molecular and Cellular Biology
|August 1, 1995
PubMed
Summary

Fission yeast possesses two DNA repair pathways for UV damage: nucleotide excision repair (NER) and a distinct SPDE-dependent pathway. The rad12-502 mutant lacks SPDE activity, confirming its role in this alternative DNA repair mechanism.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • UV radiation induces DNA lesions like cyclobutane pyrimidine dimers and (6-4)pyrimidine pyrimidones.
  • Nucleotide excision repair (NER) is a known pathway for repairing these UV-induced DNA lesions.
  • Fission yeast, Schizosaccharomyces pombe, exhibits at least two mechanisms for excising UV photoproducts, one being NER.

Purpose of the Study:

  • To investigate a previously unidentified DNA excision repair pathway in Schizosaccharomyces pombe.
  • To determine the role of the DNA endonuclease SPDE in UV damage repair.
  • To characterize the function of the rad12 gene in DNA repair.

Main Methods:

  • Utilized UV irradiation of Schizosaccharomyces pombe.
  • Assayed DNA endonuclease activity of SPDE in wild-type and rad12-502 mutant extracts.

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  • Performed in vitro DNA excision repair assays using mutant and wild-type cell extracts.
  • Generated and analyzed double mutants by crossing rad12-502 with the NER rad13-A mutant.

    • Main Results:

    • The UV-sensitive rad12-502 mutant was found to lack SPDE activity.
    • Cell extracts from the rad12-502 mutant showed deficient DNA excision repair.
    • Restoration of DNA repair activity was observed in rad12-502 extracts upon addition of purified SPDE.
    • The rad12-502 rad13-A double mutant exhibited significantly higher UV sensitivity than single mutants.

    Conclusions:

    • The rad12 gene product is essential for a DNA repair pathway distinct from NER.
    • SPDE activity is directly linked to this alternative DNA excision repair process in S. pombe.
    • A novel SPDE-dependent DNA repair pathway is proposed to exist in fission yeast.