DNA polymerase III is required for mutagenesis after damage to the chromosome. This effect is not modulated by the presence or absence of DNA polymerase II activity in the cell. In cells containing a temperature-sensitive dnaE mutation, the alpha-subunit of DNA polymerase III is inactivated at the restrictive temperature, resulting in lethality. Cells containing the pcbA1 mutation can continue replication if DNA polymerase I activity is present. When such cells are shifted from the permissive to the restrictive temperature, mutagenesis decreases rapidly after 10 min. These results are compatible with conversion of the replicative apparatus from one containing a functional DNA polymerase III synthetic subunit to one containing DNA polymerase I. We also find that DNA polymerase I dependent replication is markedly sensitive to coumermycin A1. We conclude that DNA polymerase III holoenzyme with the alpha-subunit is required for fixing mutations in the genome.