STUDIES ON THE EFFECT OF THE COMBINED EXPRESSION OF ANTI-TAT AND ANTI-REV GENES ON HIV-1 REPLICATION

A series of retroviral vectors with potential anti-tat and anti-rev ac tivity was developed. Vectors containing a tat trans-dominant negative mutant (tat(22/37)) and an RRE decoy in different positions, directed by the same promoter or by different promoters, were generated. Retro viral vectors containing tat 22/37 and the RevM10 transdominant negati ve mutant were also constructed. Jurkat cells were transduced with the recombinant retroviruses to produce monoclonal and polyclonal culture s. in these cell lines the recombinant proviruses were correctly integ rated and expression of the inserted genes was detected by Northern bl ot or RT-PCR analysis. However, infection of these cell lines with HIV -1 showed that none of these recombinant constructs inhibited virus re plication at a high multiplicity of infection (MOI). At a low MOI, two cell clones containing tat(22/37) and the RRE decoy in 3' position sh owed a long lasting protection against virus replication, in compariso n to control cultures expressing tat(22/37) or RRE alone. Combination of tat and rev mutants was ineffective in inhibiting HIV-1 replication at both low and high MOIs. At a low MOI, HIV-1 replication was effici ently blocked in two cell clones expressing the RevM10 mutant alone. T hese results show a synergic effect of anti-tat and anti-rev molecules when the RRE sequence is cloned 3' to tat(22/37), suggesting the poss ibility of using this vector design to control HIV-1 replication.