Multiple recombination events maintain sequence identity among members of the nitrogenase multigene family in Rhizobium etli

Abstract

A distinctive characteristic of the Rhizobium genome is the frequent finding of reiterated sequences, which often constitute multigene families. Interestingly, these families usually maintain a high degree of nucleotide sequence identity. It is commonly assumed that apparent gene conversion between reiterated elements might lead to concerted variation among members of a multigene family. However, the operation of this mechanism has not yet been demonstrated in the Rhizobiaceae. In this work, we employed different genetic constructions to address the role of apparent gene conversion as a homogenizing mechanism between members of the plasmid-located nitrogenase multigene family in Rhizobium etli. Our results show that a 28-bp insertion into one of the nitrogenase reiterations can be corrected by multiple recombination events, including apparent gene conversion. The correction process was dependent on the presence of both a wild-type recA gene and wild-type copies of the nitrogenase reiterations. Frequencies of apparent gene conversion to the wild-type nitrogenase reiterations were the same when the insertion to be corrected was located either in cis or in trans, indicating that this event frequently occurs through intermolecular interactions. Interestingly, a high frequency of multiple crossovers was observed, suggesting that these large plasmid molecules are engaging repeatedly in recombination events, in a situation akin to phage recombination or recombination among small, high-copy number plasmids.