ESCHERICHIA-COLI RNA AND DNA-POLYMERASE BYPASS OF DIHYDROURACIL - MUTAGENIC POTENTIAL VIA TRANSCRIPTION AND REPLICATION

Dihydrouracil (DHU) is a DNA base damage product produced in significa nt amounts by ionizing radiation damage to cytosine under anoxic condi tions. DHU represents a model for pyrimidine base damage (ring saturat ion products) of the type recognized and repaired by Escherichia coli endonuclease III and its homologs in other species. We have built this lesion into synthetic oligonucleotides, with DHU placed at a single l ocation downstream from an E. coli RNA polymerase promoter. This const ruct was used to determine the effect of DHU when encountered on a DNA template strand by either E. coli RNA or DNA polymerase (Klenow fragm ent). Single round transcription experiments or primer extension-type replication experiments were conducted in order to make a direct compa rison between RNA and DNA polymerases with DHU placed within the same sequence context. Both DNA and RNA polymerase efficiently bypass DHU a nd insert adenine opposite this lesion. These results suggest that DHU is mutagenic with respect to both replication end transcription and h ave implications for DNA repair as well the routes leading to generati on of mutant proteins in dividing and non-dividing cells.