DNA polymerase I is an efficient reverse transcriptase that mediates RNA-templated DNA repair synthesis

It is well established that during normal growth and under stress conditions ribonucleotides become nested in genomic DNA. Given the high frequency with which RNA is found in genomic DNA, it is unknown which DNA polymerase is responsible for catalyzing RNA-dependent DNA synthesis. In this work we establish bacterial DNA polymerase I (Pol I) as a robust reverse transcriptase. We tested several bacterial Pol I enzymes and found that all possess RT activity with <i>B. subtilis</i> Pol I showing RNA-dependent DNA synthesis activity similar to the Moloney Murine Leukemia Virus reverse transcriptase. We tested RNA embedded in DNA and found that replicative DNA polymerase PolC arrests at a single template ribonucleotide while, in contrast, Pol I is able to efficiently traverse long stretches of templated ribonucleotides. We conclude that Pol I is responsible for RNA templated DNA replication, allowing for replication forks to navigate RNA that persists in genomic DNA.