DNA polymerase function in repair synthesis in human fibroblasts.


We have analyzed DNA polymerase function in repair synthesis in human ...
We have analyzed DNA polymerase function in repair synthesis in human diploid fibroblasts. Previous studies from many laboratories have resulted in confusing and apparently contradictory findings; our studies suggest a unifying hypothesis and indicate that previous results may not actually be in conflict. Nongrowing (confluent) cells were damaged with N-methyl-N-nitrosourea (MNU), bleomycin, X-ray, UV radiation, or N-acetoxy-2-acetylaminofluorene (NA-AAF) over a wide range of doses, and repair synthesis was studied in the presence of one of three polymerase inhibitor, aphidicolin, dideoxythymidine triphosphate (ddTTP), or N-ethyl maleimide. We find that both polymerase alpha and a non-alpha polymerase, probably polymerase beta, are involved in repair synthesis. Furthermore, there is a dose dependence for polymerase function: at low doses of damage repair synthesis is mediated primarily by the non-alpha polymerase; as the amount of damage is increased, polymerase alpha participates to an increasing extent and at high levels of damage is responsible for approximately 50-80% of the repair synthesis. In a study in rapidly growing cells, repair synthesis following a high dose of UV was refractory to aphidicolin; these findings indicate that in growing and quiescent cells DNA polymerases may function differently in repair synthesis. All of the above results may be analyzed in terms of a simple model in which two repair synthesis systems, each involving one of the polymerases, compete to put repair patches into damaged DNA.




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