Stable and specific association between the yeast recombination and DNA repair proteins RAD1 and RAD10 in vitro.
The RAD1 and RAD10 genes of Saccharomyces cerevisiae are two of at least seven genes which are known to be required for damage-specific recognition and/or damage-specific incision of DNA during nucleotide excision repair. RAD1 and RAD10 are also involved in a specialized mitotic recombination pathway. We have previously reported the purification of the RAD10 protein to homogeneity (L. Bardwell, H. Burtscher, W. A. Weiss, C. M. Nicolet, and E. C. Friedberg, Biochemistry 29:3119-3126, 1990). In the present studies we show that the RAD1 protein, produced by in vitro transcription and translation of the cloned gene, specifically coimmunoprecipitates with the RAD10 protein translated in vitro or purified from yeast. Conversely, in vitro-translated RAD10 protein specifically coimmunoprecipitates with the RAD1 protein. The sites of this stable and specific interaction have been mapped to the C-terminal regions of both polypeptides. This portion of RAD10 protein is evolutionarily conserved. These results are the first biochemical evidence of a specific association between any eukaryotic proteins genetically identified as belonging to a recombination or DNA repair pathway and suggest that the RAD1 and RAD10 proteins act at the same or consecutive biochemical steps in both nucleotide excision repair and mitotic recombination.
Bardwell
L
L
Department of Pathology, University of Texas Southwestern Medical Center, Dallas 75235.
Cooper
A J
AJ
Friedberg
E C
EC
D001483:Base Sequence; D001665:Binding Sites; D002874:Chromosome Mapping; D004252:DNA Mutational Analysis; D004260:DNA Repair; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins; D004720:Endonucleases; D005656:Fungal Proteins / Q000378:metabolism*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D015719:Single-Strand Specific DNA and RNA Endonucleases
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D029701:Saccharomyces cerevisiae Proteins; D004720:Endonucleases; C069266:RAD1 protein, S cerevisiae; C064162:RAD10 protein, S cerevisiae; D015719:Single-Strand Specific DNA and RNA Endonucleases; D045643:DNA Repair Enzymes