MUTATIONAL ANALYSIS OF ERCC3, WHICH IS INVOLVED IN DNA-REPAIR AND TRANSCRIPTION INITIATION - IDENTIFICATION OF DOMAINS ESSENTIAL FOR THE DNA-REPAIR FUNCTION

The human ERCC3 gene, which corrects specifically the nucleotide excis ion repair defect in human xeroderma pigmentosum group B and cross-com plements the repair deficiency in rodent UV-sensitive mutants of group 3, encodes a presumed DNA helicase that is identical to the p89 subun it of the general transcription factor TFIIH/BTF2. To examine the sign ificance of the postulated functional domains in ERCC3, we have introd uced mutations in the ERCC3 cDNA by means of site-specific mutagenesis and have determined the repair capacity of each mutant to complement the W-sensitive phenotype of rodent group 3 cells. A conservative subs titution of arginine for the invariant lysine residue in the ATPase mo tif (helicase domain I), six deletion mutations in the other helicase domains, and a deletion in the potential helix-turn-helix DNA-binding motif fail to complement the ERCC3 excision repair defect of rodent gr oup 3 mutants, which implies that the helicase domains as well as the potential DNA-binding motif are required for the repair function of ER CC3. Analysis of carboxy-terminal deletions suggests that the carboxy- terminal exon may comprise a distinct determinant for the DNA repair f unction. In addition, we show that a functional epitope-tagged version of ERCC3 accumulates in the nucleus. Deletion of the putative nuclear location signal impairs neither the nuclear location nor the repair f unction, indicating that other sequences may (also) be involved in tra nslocation of ERCC3 to the nucleus.