| All organisms need to replicate their genomes accurately to survive. The bacterium Escherichia coli has five known DNA polymerases. DNA Polymerase IV (Pol IV), is characterized by its high frequency of nucleotide misincorporation, its production of frameshift mutations, and its ability to replicate damaged DNA. Although Pol IV is not essential for cell viability, it may help to rescue replication forks when the replicative DNA polymerase, Pol III, stalls at DNA lesions. Pol IV is highly expressed and has the ability to polymerize hundreds of nucleotides using an undamaged template. However, Pol IV's mutational activity appears to be restricted to conditions of cellular stress. Therefore, we hypothesized that under normal conditions Pol IV activity is controlled by restricting its access to the replication fork. To test this hypothesis, I used a genetic approach to investigate what cellular factors and conditions regulate Pol IV. First, I provide support for a model that Pol IV-dependent adaptive mutations that occur during nutrient limitation are most likely due to highly mutagenic recombination-dependent replication. Second, I demonstrate that Pol IV is regulated by polyphosphate kinase (Ppk) and the chains of inorganic polyphosphate (polyP) that Ppk synthesizes. Finally, I show that a previously unknown small regulatory RNA (spk) suppresses the loss of Ppk and restores Pol IV activity. These studies provide the first examples of regulatory factors that affect Pol IV's activity without changing its cellular abundance, suggesting that Ppk and spk regulate Pol IV's mutagenic activity, or its access to the replication fork, even under non-stressful conditions. |
