Induction of a mutant phenotype in human repair proficient cells after overexpression of a mutated human DNA repair gene.
Antisense and mutated cDNA of the human excision repair gene ERCC-1 were overexpressed in repair proficient HeLa cells by means of an Epstein-Barr-virus derived cDNA expression vector. Whereas antisense RNA did not influence the survival of the transfected cells, a mutated cDNA generating an ERCC-1 protein with two extra amino acids in a conserved region of its C-terminal part resulted in a significant sensitization of the HeLa transfectants to mitomycin C-induced damage. These results suggest that overexpression of the mutated ERCC-1 protein interferes with proper functioning of the excision repair pathway in repair proficient cells and is compatible with a model in which the mutated ERCC-1 protein competes with the wild-type polypeptide for a specific step in the repair process or for occupation of a site in a repair complex. Apparently, this effect is more pronounced for mitomycin C induced crosslink repair than for UV-induced DNA damage.
Belt
P B
PB
Department of Biochemistry, Gorlaeus Laboratories, Leiden University, The Netherlands.
van Oosterwijk
M F
MF
Odijk
H
H
Hoeijmakers
J H
JH
Backendorf
C
C
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins; D004720:Endonucleases; D005786:Gene Expression Regulation; D006367:HeLa Cells / Q000187:drug effects / Q000528:radiation effects; D006801:Humans; D051379:Mice; D016685:Mitomycin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D011506:Proteins / Q000235:genetics*; D016372:RNA, Antisense / Q000378:metabolism; D016415:Sequence Alignment; D014162:Transfection; D014466:Ultraviolet Rays
D004268:DNA-Binding Proteins; D011506:Proteins; D016372:RNA, Antisense; D016685:Mitomycin; C071447:ERCC1 protein, human; D004720:Endonucleases; C499360:Ercc1 protein, mouse