RNA motifs mediating in vivo site-specific nonhomologous recombination in (+) RNA virus enforce in vitro nonhomologous crossovers with HIV-1 reverse transcriptase

There are several lines of evidence that both RNA viruses and retroviruses recombine according to a copy choice mechanism. Using the brome mosaic viru s (BMV)-based system, we recognized elements in the RNA structure that enha nce nonhomologous crossovers within or near the local heteroduplex formed b y recombining molecules. The same structural motifs were employed in vitro to test the ability of human immunodeficiency virus reverse transcriptase ( HIV-RT) to switch templates during DNA synthesis. We demonstrated that a sp ecific combination of the local double-stranded region with short homologou s sequences and a hairpin structure allows template switching by HIV-RT. In contrast to BMV replicase, HIV-RT does not mediate the detectable level of recombination using only the heteroduplex structure, though local hybridiz ation between RNA molecules efficiently pauses primer extension. Moreover, the presented data suggest that a proper arrangement of identified structur al motifs can ensure site specificity of RNA-RNA recombination. These resul ts indicate that HIV-RT utilizes the same or a very similar mechanism as BM V replicase to change nonhomologous RNA templates in a site-specific manner .