Dimerization of HIV-1 genomic RNA of subtypes A and B: RNA loop structure and magnesium binding

Retroviruses encapsidate their genome as a dimer of homologous RNA molecule s noncovalently linked close to their 5' ends. The dimerization initiation site (DIS) of human immunodeficiency virus type 1 (HIV-1) RNA is a hairpin structure that contains in the loop a 6-nt self-complementary sequence flan ked by two 5' and one 3' purines. The self-complementary sequence, as well as the flanking purines, are crucial for dimerization of HIV-1 RNA, which i s mediated by formation of a "kissing-loop" complex between the DIS of each monomer. Here, we used chemical modification interference, lead-induced cl eavage, and three-dimensional modeling to compare dimerization of subtype A and B HIV-1 RNAs. The DIS loop sequences of these RNAs are AGGUGCACA and A AGCGCGCA, respectively. In both RNAs, ethylation of most but not all phosph ate groups in the loop and methylation of the N7 position of the G residues in the self-complementary sequence inhibited dimerization. These results d emonstrate that small perturbations of the loop structure are detrimental t o dimerization. Conversely, methylation of the N1 position of the first and last As in the loop were neutral or enhanced dimerization,a result consist ent with these residues forming a noncanonical sheared base pair. Phosphoro thioate interference, lead-induced cleavage, and Brownian-dynamics simulati on revealed an unexpected difference in the dimerization mechanism of these RNAs. Unlike subtype B, subtype A requires binding of a divalent cation in the loop to promote RNA dimerization. This difference should be taken into consideration in the design of antidimerization molecules aimed at inhibit ing HIV-1 replication.