[PubMed] [Google Scholar] 42. Rev transactivator activity. Consistent with these results, a HIV-1 proviral plasmid that expresses a C-terminally truncated Rev mutant protein produces smaller amounts of the p24 antigen than does a plasmid that possesses an undamaged gene. These results indicate the practical importance of the C-terminal region for full Rev activity, which leads to efficient HIV-1 replication. Human being immunodeficiency disease type 1 (HIV-1) encodes the Rev protein, which localizes mainly to the nucleus and/or nucleolus (4, 5, 10, 27) and actively shuttles between the nucleus and PFI-3 cytoplasm (35, 45, 50). Rev exports unspliced or incompletely spliced viral mRNAs to the cytoplasm, therefore assisting the efficient manifestation of the late viral enzymatic and structural proteins encoded by these mRNAs (9, 11, 14, 15, 16, 25, 52). To day, three practical domains indispensable for Rev function have been identified. One is an arginine-rich fundamental domain located in the region of amino acids (aa) 35 to 50 of Rev, which has been shown to mediate the direct nuclear binding of Rev to the highly organized DNA polymerase to produce blunt ends, digested with polymerase, and coding region from pSRRev or pM10 (42) was amplified by PCR using the primer pair primer 1 and 5-GGA ATT CTA AGC GTA GTC TGG GAC GTC GTA TGG GTA TTC TTT AGC TCC TGA CTC CA-3, which encodes a hemagglutinin (HA) tag. The PCR products were digested with sequences were amplified by PCR using pSRRev like a template and the primer pair 5-CTG CAA AGC TTA TGG CAG GAA GAA GCG GAG-3 (primer 4) and primer 2 or primer 4 and primer 3, respectively. Each PCR product was cloned into pSGGALVP (20), which had been treated with polymerase and was then digested with sequence was amplified by PCR using pSRRevM10-HA like a template and the primer pair primer 4 and 5-GAG GTA CCT ATT ATT TAG CTC CTG Take action C-3 (primer 5). The PCR product was cloned into pSGGALVP as explained above. To make pRev86-VP or pRev100-VP, a corresponding region of PFI-3 the sequence was amplified by PCR using pSRRev like a template and the primer pair Mcam 5-GCC ATG GCA GGA AGA AGC GGA G-3 (primer 6) and 5-GGC CCA AGC TTG TTA CAA TCA AGA GTA AGT CT-3 or primer 6 and 5-GGC CCA AGC TTA GGG CTT CCC ACC CCC TGC GT-3, respectively. Each PCR product PFI-3 was cloned into pSGGALVP, which had been treated sequentially with polymerase, and sequence was amplified by PCR using pSRRevM10-HA like a template and the primer pair 5-CCG AAT TCA CCA TGG CAG GAA GAA GCG GAG A-3 and 5-TTC CCA AGC TTT TCT TTA GCT CCT GAC TCC A-3. The PCR product was cloned into pSGGALVP as explained above. The plasmids pSRhCRM1, pGAL-hCRM1, pGAL4, pG5BCAT, pG5BLuc (23, 24), pSRRev, pGAL-Rev, pRev-VP, pGAL-Rex, pRex-VP, pCDM-galactosidase (pCDM-Gal) (36), pGAL-RXR, pRAR-VP (18), and pDM128 (32) have been previously explained. The recombinant RanQ69L manifestation plasmid pET3dRanQ69L and recombinant hCRM1 and the zz-Rev protein manifestation plasmids pET3hCRM1 and p6z60Rev were kind gifts from Y. Yoneda (pET3dRanQ69L) and D. G?rlich (hCRM1 and remaining plasmids) (29, 49). pNL4-3.Luc.E?R? (7), which is derived from the HIV-1 proviral clone pNL4-3, is referred to as pNL4-3LucE?R?(Rev) with this study. To construct pNL4-3LucE?R?(Rev86), which expresses Rev86, PCR-based site-directed mutagenesis using pNL4-3LucE?R?(Rev) like a template and the primer pair 5-CTC TTG ATT GTA ACG AGG ATT GTG GAA CTT CTG GGA CG-3 and its complementary oligonucleotide was done. The insertion of PFI-3 a termination codon to delete the C-terminal region (aa 87 to 116) of Rev does not affect some other viral PFI-3 protein.