Expression of biologically active HIV glycoproteins using a T7 RNA polymerase-based eucaryotic vector system.
Bacteriophage T7 RNA polymerase and a derivative containing a nuclear localization signal were transiently expressed in CV-1 cells and were shown to localize to the cytoplasm and nucleus, respectively. A vector was constructed containing T7 promoter and transcription terminator sequences flanking a picornaviral 5' untranslated sequence for cap-independent translation and a polyA signal. Expression of the HIV-1 envelope glycoproteins in this vector system gave high levels of specific transcripts and translation products, independent of the subcellular localization of T7 RNA polymerase. The synthesis of HIV glycoproteins was also completely independent of the coexpression of the HIV rev protein, which is normally required for the expression of HIV structural proteins. In addition, a polyA signal was not required, whereas the presence of the picornaviral 5' untranslated region was necessary for efficient expression. Different possibilities to account for these findings are discussed. The HIV glycoproteins synthesized in this system were normally processed and assembled; they could induce syncytium formation and complement an env-deletion mutant of HIV-1.
Wilk
T
T
Institut für Virusforschung, Deutsches Krebsforschungszentrum, Heidelberg, FRG.
Mierswa
H
H
Kräusslich
H G
HG
Dunn
J J
JJ
Bosch
V
V
D000818:Animals; D017123:Bacteriophage T7; D002460:Cell Line; D012321:DNA-Directed RNA Polymerases; D015870:Gene Expression / Q000235:genetics; D015752:Genes, env / Q000502:physiology*; D015757:Genes, rev / Q000502:physiology; D005822:Genetic Vectors; D006023:Glycoproteins / Q000096:biosynthesis*; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014764:Viral Proteins / Q000096:biosynthesis*
D006023:Glycoproteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases