Low fidelity DNA polymerase IV accelerates genome evolution in Pseudomonas aeruginosa

Specialized DNA polymerases are crucial for bypassing lesions and facilitating various cellular processes. Despite extensive research, the mutagenic effects of these error-prone enzymes on genomes are still not fully understood. In this study, we examined the genomic instability caused by Pol IV in the bacterial pathogen Pseudomonas aeruginosa. Pol IV mutagenesis primarily involved the misincorporation of oxidized guanine nucleotides opposite template adenine. This activity led to a distinctive mutational signature, characterized by A to C transversions occurring preferentially at AT sites flanked by a 5G and/or 3C. Furthermore, Pol IV preferentially targeted specific chromosomal locations near the replication termination region and rRNA-encoding operons. Several genes associated with virulence, motility, antibiotic resistance and chemotaxis are located in these difficult-to-replicate regions, making them hotspots for Pol IV-mutagenesis. Notably, half of the mutation events catalyzed by Pol IV effectively impaired gene function. This can be attributed to the strong bias of Pol IV for mutating specific codons with its preferred sequence contexts, leading to substitutions primarily to the unreactive Ala and Gly residues. Remarkably, mutation signatures identified for Pol IV were also present in genomes from clinical isolates of P. aeruginosa, providing compelling evidence for its role in genetic diversification during pathogen adaptation.