We have comprehensively analyzed the distribution of functionally critical sequence-specific information throughout the RRE, the target seguence for the REV protein of human immunodeficiency virus (HIV-1). We found that seguence-specific information is widespread throughout the RRE, both within and without the primary REV binding domain. The finding of sequence-specific information outside the primary REV binding domain suggests that this region may be specifically bound by cellular factors involved in the REV response. Further we found that sequence-specific information is about equally distributed amongst the stems of the REV binding doman. The most critical sequence-specific information in the entire molecule lies at the center of the REV binding domain, in llb. Sequence specificity here is clearly found on both strands of the RNA and was identified at the level of single bases. This region may act as a nucleation center for REV protein polymerization along the RNA molecule or may be involved in the specific binding of cellular factors. Using this information, and information we previously generated on the structure of the RRE, we have designed a series of RNA probes to study the binding of REV to the RRE and to identify cellular factors involved in the REV response. Initial studies have detected a nuclear activity which promotes REV binding to the RRE. Studies are now underway to study the specificity of this interaction and to characterize, identify and clone the nuclear factor(s) involved. In other work, we have initiated studies on the possible interaction of the vif protein of HIV-1 with other viral genes. HIV clones with vif mutations in different viral genetic backgrounds have different cellular tropisms which do not clearly mirror parental tropisms. Exchanging sequences between two vif mutants with divergent tropisms determined that some of the sequences responsible for the differences lie in a large envelope-containing region of the genome. We have also accumulated evidence that the RRE can contribute to cell-type specific viral tropism.
