CHARACTERIZATION OF A MAMMALIAN REPAIR GENE

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DESCRIPTION:Although it is known that normal mammalian cells employ mutagenic replicative bypass as a tolerance mechanism for certain DNA lesions, much remains to be learned about this pathway of DNA repair/damage tolerance at the molecular level. A CHO mutant, called CHO-UV1, showing defective mutagenic bypass repair has been isolated. UV1 is hypersensitive to killing by various DNA damaging agents including alkylating drugs, mitomycin C and UV, but not ionizing radiation. UV1 has normal nucleotide excision repair but is defective in bypass repair and post replication recovery (PRR). It is less mutable than wild type CHO implying that the normal UV1 "gene" function is mutagenic. The phenotype of UV1 can be complemented by transfecting human genomic DNA into UV1 or by fusing UV1 with normal human cells, suggesting that human cells possess an equivalent of the gene mutated in UV1. A human cDNA encoding a truncated form of the ALY transcriptional coactivator has been found to complement the MMC killing hypersensitivity of UV1. Antisense-ALY, on the other hand, increases CHO's sensitivity to MMC. A fully MMC-resistant human-UV1 hybrid has been found to contain a fragment of human chromosome 11p with the ALY gene in it. The preliminary evidence thus implicates ALY as a candidate for the UV1 "gene" and a role for ALY in bypass repair and PRR. Studies are proposed to: (1) Investigate in detail the role of ALY in the UV1 PRR repair pathway, (2) examine the possibility that another gene can complement the UV1-PRR pathway and further characterize the phenotype of the mutant, and (3) examine the biochemical role of ALY (and/or other complementing genes) in the mutagenic PRR process.