YEAST RAD7-RAD16 COMPLEX, SPECIFIC FOR THE NUCLEOTIDE EXCISION-REPAIROF THE NONTRANSCRIBED DNA STRAND, IS AN ATP-DEPENDENT DNA-DAMAGE SENSOR

In eukaryotes, nucleotide excision repair of ultraviolet light-damaged DNA is a highly intricate process that requires a large number of evo lutionarily conserved protein factors, Genetic studies in the yeast Sa ccharomyces cerevisiae have indicated a specific role of the RAD7 and RAD16 genes in the repair of transcriptionally inactive DNA. Here we s how that the RAD7- and RAD16-encoded products exist as a complex of 1: 1 stoichiometry, exhibiting an apparent dissociation constant (K-d) of <4 x 10(-10) M. The Rad7-Rad16 complex has been purified to near homo geneity in this study and is shown to bind, in an ATP-dependent manner and with high specificity, to DNA damaged by ultraviolet light, Impor tantly, inclusion of the Rad7-Rad16 complex in the in vitro nucleotide excision repair system that consists entirely of purified components results in a marked stimulation of damage specific incision. Thus, Rad 7-Rad16 complex is the ATP-dependent DNA damage sensor that specifical ly functions with the ensemble of nucleotide excision repair factor (N EF) 1, NEF2, NEF3, and replication protein A in the repair of transcri ptionally inactive DNA. We name this novel complex of Rad7 and Rad16 p roteins NEF4.