SCANNING MUTAGENESIS OF THE ARGININE-RICH REGION OF THE HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 REV TRANS ACTIVATOR

The structural proteins of human immunodeficiency virus type 1, for ex ample, Gag and Env, are encoded by unspliced and incompletely spliced viral transcripts. The expression of these mRNAs in the cytoplasm, alo ng with their commensurate translation, is absolutely dependent on the virally encoded Rev trans activator. Previous studies have demonstrat ed that Rev binds directly to its substrate mRNAs via an arginine-rich element that also serves as its nuclear localization sequence. In an attempt to define the specific amino acid residues that are important for in vivo activity, we have constructed a series of missense mutatio ns that scan across this region. Our data demonstrate that all eight a rginine residues within this element can, individually, be substituted for either leucine or lysine with no apparent loss of function. Impor tantly, these findings suggest that no single amino acid within the ar ginine-rich domain of Rev is, by itself, essential for activity and th at considerable functional redundancy is therefore likely to exist wit hin this region. Interestingly, one mutant in which a tryptophan had b een substituted for a serine failed to accumulate exclusively in the n ucleus but still bound RNA in a manner that was indistinguishable from that of the wild-type protein. This observation indicates that featur es of the arginine-rich region that are additional to those required f or RNA binding are important for Rev's correct accumulation in the nuc leus.