INTERACTION WITH THE REV RESPONSE ELEMENT ALONG AN EXTENDED STEM-I DUPLEX STRUCTURE IS REQUIRED TO COMPLETE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 REV-MEDIATED TRANSACTIVATION IN-VIVO

Expression of structural proteins of human immunodeficiency virus type 1 requires the direct interaction of the viral Rev traits-activator with its cis-acting RNA target sequence, the Rev response element (RRE). The originally defined 234-residue RRE, however, failed to show the full HIV-1 regulator of virion expression (Rev) response when inserted into a test plasmid that lacked the flanking env sequences. Here, we demonstrate that an alternative 351-residue complete RRE, which carries an extended Stem I structure, is required for the full Rev-responsiveness in vivo. Mutagenesis studies revealed that the association of Rev with RRE was initiated by the recognition of a functional bubble structure in stem-loop II, followed by interaction with the neighbouring duplex RNA, including the elongated Stem I structure. We propose that, as has been previously shown in vitro, the Rev-RRE interaction in vivo is an ordered assembly process and is based on the association of Rev molecules with the double-stranded region of RRE.