Human immunodeficiency virus type-1 reverse transcription can be inhibitedin vitro by oligonucleotides that target both natural and synthetic tRNA primers

Reverse transcription of human immunodeficiency virus type-1 is primed by c ellular tRNA(Lys3), which is selectively packaged into viral particles wher e it is bound at its 3' terminus to a complementary sequence of viral RNA t ermed the primer binding site (PBS). Since cellular tRNA(Lys3) is highly co nserved, it might conceivably serve as a good target for novel antagonists to block reverse transcriptase (RT) activity. In this study, we have examin ed a number of antisense oligodeoxyribonucleotides (ODNs) that are compleme ntary to different parts of the tRNA primer and, therefore, may interfere w ith the initiation of RT-mediated DNA synthesis. We found that the stabilit y of complexes between synthetic tRNA(Lys3) and ODNs was significantly incr eased when binding occurred via sequences involved in tertiary interactions of the tRNA. In particular, ODNs with complementarity to both the variable and T Psi C stem-loop of tRNA(Lys3) bound with high affinity to both free tRNA(Lys3) as well as to the binary tRNA(Lys3)/RNA complex. As a result, th e initiation of DNA synthesis was severely compromised under these conditio ns. Moreover, RT-associated RNase H activity recognized the tRNA within thi s ternary tRNA(Lys3)/RNA/ODN complex as an RNA template and initiated its d egradation. Both this RNase H degradation of tRNA(Lys3) as well as the alte red structure of the tRNA/RNA complex, due to the binding of the ODN, contr ibuted to the inhibition of synthesis of viral DNA. The initiation of RT ac tivity was almost completely blocked when using ODNs that interfered with i ntermolecular tRNA/RNA interactions that involved both the PBS and sequence s outside the PBS, Similar findings were obtained with natural preparation of tRNA(Lys3).