R hodobacter capsulatus DprA is essential for RecA ‐mediated gene transfer agent ( RcGTA ) recipient capability regulated by quorum‐sensing and the CtrA response regulator

Summary Gene transfer agents ( GTAs ) are genetic exchange elements that resemble small DNA bacteriophages that transfer random pieces of the producing cell's genome to recipient cells. The best‐studied GTA is that of R hodobacter capsulatus , termed RcGTA . We discovered that the putative response regulator CtrA , which is essential for RcGTA production, is required for RcGTA ‐mediated gene acquisition, and confirmed that a RecA homologue is required. It was also discovered that a DprA ( DNA ‐protecting protein A ) homologue is essential for RcGTA ‐mediated gene acquisition, and that dprA expression is induced by gtaI ‐dependent quorum‐sensing and non‐phosphorylated CtrA . Modelling of the R . capsulatus   DprA structure indicated the presence of a C ‐terminal region that resembles a dsDNA ‐binding protein domain. Purified His‐tagged R . capsulatus   DprA protein bound to both single‐stranded (ss) DNA and double‐stranded (ds) DNA , but with a greater affinity for ssDNA . Additionally, DprA protected dsDNA from endonuclease digestion, and increased the rate of nucleation of E scherichia coli   RecA onto ssDNA . Single‐cell expression analyses revealed that dprA is expressed in the majority of cells throughout a population. Overall, the results suggest that incorporation of RcGTA DNA into the recipient cell genome proceeds through a homologous recombination pathway resembling DNA recombination in natural transformation.