FUNCTIONAL ANALYSIS OF RAD23 PROTEIN

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We have discovered that the DNA repair protein Rad23 has a proteolytic function. The studies proposed here have broad relevance to radiation biology, and can make a strong contribution to a molecular understanding of nucleotide excision-repair from yeast to humans. The ubiquitin/proteosome pathway is required for the degradation of many key regulators in eukaryotic cells. The targets of this pathway include important cell-cycle regulators, transcription factors, tumor suppressors, and signal-transducing molecules. Our preliminary findings revealed a role for protein degradation in nucleotide excision-repair; i) we found that Rad23 has a proteolytic function. ii) Rad 23 is phosphorylated by the Rad53 kinase, which mediates a DNA damage-inducible checkpoint. iii) Specific ubiquitin-conjugating (E2) enzymes, including the DNA repair protein Rad6, interact with the proteosome. These studies provide a strong foundation for studying the role of protein degradation in DNA repair. The questions that are addressed in this Grant Application can yield new and interesting information on the proteolytic function of Rad23. These studies can also increase our understanding of the mechanisms of substrate targeting, and the significance of E2/proteasome interaction. The consequence of Rad23/proteasome interaction in DNA repair will be investigated. Our studies have added significance because the human homologs of Rad23 are functionally similar to their yeast counterpart. An underlying defect in several human diseases, including xeroderma pigmentosum and Cockayne's syndrome, involves the nucleotide excision-repair pathway. The present understanding of nucleotide excision-repair and ubiquitin-mediated proteolysis is well developed, and we now have the ability to form a coherent model that describes the link between these two pathways.