A loop-loop "kissing" complex is the essential part of the dimer linkage of genomic HIV-1 RNA.

Abstract

RNA-RNA interactions govern a number of biological processes. Several RNAs, including natural sense and antisense RNAs, interact by means of a two-step mechanism: recognition is mediated by a loop-loop complex, which is then stabilized by formation of an extended intermolecular duplex. It was proposed that the same mechanism holds for dimerization of the genomic RNA of human immunodeficiency virus type 1 (HIV-1), an event thought to control crucial steps of HIV-1 replication. However, whereas interaction between the partially self-complementary loop of the dimerization initiation site (DIS) of each monomer is well established, formation of the extended duplex remained speculative. Here we first show that in vitro dimerization of HIV-1 RNA is a specific process, not resulting from simple annealing of denatured molecules. Next we used mutants of the DIS to test the formation of the extended duplex. Four pairs of transcomplementary mutants were designed in such a way that all pairs can form the loop-loop "kissing" complex, but only two of them can potentially form the extended duplex. All pairs of mutants form heterodimers whose thermal stability, dissociation constant, and dynamics were analyzed. Taken together, our results indicate that, in contrast with the interactions between natural sense and antisense RNAs, no extended duplex is formed during dimerization of HIV-1 RNA. We also showed that 55-mer sense RNAs containing the DIS are able to interfere with the preformed HIV-1 RNA dimer.

The bifurcated stem loop 4 (SL4) is crucial for efficient packaging of mouse mammary tumor virus (MMTV) genomic RNA

“…Thus, in MMTV, a combination of both cis -acting sequences and structural elements are responsible for the recognition and differential packaging of the unspliced RNA during the encapsidation process. The case of human immunodeficiency virus type 1 (HIV-1) may somewhat be similar to MMTV: the 5ʹ UTR in HIV-1 implicated in RNA packaging folds into several stem loops, SL1-4 [39,40] of which SL1 harbors a GC-rich 6-nt pal sequence in the form of a loop that has been shown to function as the DIS [41,42]. Until recently SL3, which is present downstream of mSD, had been proposed to be the main HIV-1 packaging signal since it harbors a conserved region containing a GGAG tetraloop that has been shown to bind NC [43].…”

Specific recognition of the HIV-1 genomic RNA by the Gag precursor

“…The two substitutions that were introduced in the SL1 apical loop are also different (Fig. 5a), but they both impair gRNA dimerization 33,50 . Finally, a complete deletion of SL3 was introduced in the RNAs derived from both the MAL and the NL4-3 isolates (Fig.…”

“…4a). Substitutions were introduced in the apical loop of each hairpin: a point substitution preventing RNA dimerization 50 was introduced into SL1, a 'GNRA' loop was substituted for the SL2 loop and stable 'UNCG' loops were substituted for the purine-rich SL3 and SL4 loops (Fig. 4b).…”

In vitro selection of RNAs that bind to the human immunodeficiency virus type-1 gag polyprotein

“…Different structures and sequences within ψ may be required for distinct dimer formation, gag binding and RNA packaging steps. Guanine quartets (33) and kissing loops (34)(35)(36)(37)(38)(39) have been proposed to be important for some of these steps. We considered whether the nucleocapsid-binding RNAs described here could form such structures.…”