STRUCTURAL AND FUNCTIONAL-CHARACTERIZATION OF REV-LIKE TRANSCRIPTS OFEQUINE INFECTIOUS-ANEMIA VIRUS

Three cDNA clones representing structurally distinct transcripts were isolated from a cDNA library prepared from cells infected with equine infectious anemia virus (EIAV) by using a probe representing the S3 op en reading frame, which is thought to encode Rev. One species, designa ted p2/2, contained four exons and was identical to a previously descr ibed polycistronic mRNA that encodes Tat. This transcript was predicte d to also direct the synthesis of a truncated form of the transmembran e protein and a putative Rev protein whose N-terminal 29 amino acids, derived from env, are linked to S3 sequences. The second cDNA, p176, a lso consisted of four exons which were generated by two of three of th e same splicing events that occur with p2/2 but not with the Tat mRNA. The alternative splice site giving rise to the second exon of p176 re sults in a bicistronic message that would encode the same transmembran e and Rev proteins as p2/2. The first exon of the third transcript, p2 0, was identical to those of p2/2 and p176 but was spliced directly to S3. This monocistronic message could encode a second form of Rev that lacks env sequences, provided that Rev synthesis would initiate at a non-AUG codon. The coding capacity of each cDNA was assessed in a euka ryotic system using S3 antisera. Two putative Rev proteins with appare nt molecular masses of 18 and 16 kDa were expressed by p2/2 and p176, while p20 expressed only a 16-kDa species. Analysis of EIAV-infected c ells with S3 antisera revealed the presence of an 18-kDa protein. Surp risingly, the same protein was detected in purified virions. By using a reporter construct, the chloramphenicol acetyltransferase gene linke d to EIAV env sequences, we were able to demonstrate greatly enhanced chloramphenicol acetyltransferase activity in cells cotransfected with this construct and any of the three cDNAs.