Highly mutagenic replication by DNA polymerase V (UmuC) provides a mechanistic basis for SOS untargeted mutagenesis

When challenged by DNA-damaging agents, Escherichia coil cells respond by i nducing the SOS stress response, which leads to an increase in mutation fre quency by two mechanisms: translesion replication, a process that causes mu tations because of misinsertion opposite the lesions, and an inducible muta tor activity, which acts at undamaged sites. Here we report that DNA polyme rase V (pol V; UmuC), which previously has been shown to be a lesion-bypass DNA polymerase, was highly mutagenic during in vitro gap-filling replicati on of a gapped plasmid carrying the cro reporter gene. This reaction requir ed, in addition to pol V, UmuD', RecA, and single-stranded DNA (ssDNA)-bind ing protein, pol V produced point mutations at a frequency of 2.1 x 10(-4) per nucleotide (2.1% per cro gene), 41-fold higher than DNA polymerase III holoenzyme. The mutational spectrum of pol V was dominated by transversions (53%), which were formed at a frequency of 1.3 x 10(-4) per nucleotide (1. 1% per cro gene), 74-fold higher than with pol III holoenzyme. The prevalen ce of transversions and the protein requirements of this system are similar to those of in vivo untargeted mutagenesis (SOS mutator activity). This fi nding suggests that replication by pol V, in the presence of UmuD', RecA, a nd ssDNA-binding protein, is the basis of chromosomal SOS untargeted mutage nesis.