Evidence for a Mechanism of Recombination during Reverse Transcription Dependent on the Structure of the Acceptor RNA

Genetic recombination is a major force driving retroviral evolution. In retroviruses, recombination proceeds mostly through copy choice during reverse transcription. Using a reconstituted in vitro system, we have studied the mechanism of strand transfer on a major recombination hot spot we previously identified within the genome of HIV-1. We show that on this model sequence the frequency of copy choice is strongly influenced by the folding of the RNA template, namely by the presence of a stable hairpin. This structure must be specifically present on the acceptor template. We previously proposed that strand transfer follows a two-step process: docking of the nascent DNA onto the acceptor RNA and strand invasion. The frequency of recombination under copy choice conditions was not dependent on the concentration of the acceptor RNA, in contrast with strand transfer occurring at strong arrests of reverse transcription. During copy choice strand transfer, the docking step is not rate limiting. We propose that the hairpin present on the acceptor RNA could mediate strand transfer following a mechanism reminiscent of branch migration during DNA recombination.