We determined the structure of CAR, the RNA target sequence of the rev protein of HIV-1 and identified which of the sub-structures of CAR are essential for function. Site-directed mutagenesis was employed to alter selected sequences in CAR enabling us to determine 14 regions which basepaired to form 7 stem structures. By computer analyses we will then predict with confidence the secondary structure of the entire sequence. We have continued this analysis and have accumulated evidence for limited sequence specificity in the interaction between CAR and viral and/or cellular binding proteins. Using band shift analysis we have studied the binding of REV to CAR and have tentatively identified regions which bind REV and regions which probably bind cellular factors, We have also initiated studies on the interaction of nuclear dsRNA unwindase (/adenosine deaminase) with CAR. 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. We are presently exchanging sequences between two vif mutants with divergent tropisms to determine the sequences responsible for the differences.
