Retrovirus vector-mediated transfer of functional HIV-1 regulatory genes.
Replication of the human immunodeficiency virus depends on the expression of its regulatory genes. We have constructed three plasmids, based on the retrovirus vector LXSN, that contain the tat, rev, and env (pLTRESN), the rev and env (pLRESN), and the nef (pLnefSN) genes of HIV-1. In a two-step virus rescue protocol, during which introns are removed from the DNA fragments inserted into pLXSN, these plasmids were used to establish amphotropic retrovirus vector producer lines for the transfer of tat (LtatSN), rev (LrevSN), and nef (LnefSN). These vectors have titers greater or equal to 10(6) CFU/ml and efficiently transduced each of these genes into a variety of human and murine cell lines. Representative populations of cells constitutively expressing the tat and rev genes were obtained. Cell lines transduced with LtatSN were able to trans-activate an HIV-LTRCAT construct, indicating the presence of a functional Tat protein. Similarly, cells transduced with LrevSN were able to rescue a rev- HIV-1 provirus, indicating the presence of a functional Rev. We also used LnefSN to obtain clones of cells expressing Nef. Our results indicate that these retrovirus vectors are useful reagents for the efficient transfer of functional Tat, Rev, and Nef and for the establishment of cell lines constitutively expressing these genes.
Garcia
J V
JV
Department of Virology and Molecular Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105.
Miller
A D
AD
D016475:3T3 Cells; D000818:Animals; D002460:Cell Line; D018014:Gene Transfer Techniques; D005809:Genes, Regulator; D005814:Genes, Viral; D015752:Genes, env; D015754:Genes, nef; D015757:Genes, rev; D015756:Genes, tat; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D010957:Plasmids; D012190:Retroviridae / Q000235:genetics*; D015533:Transcriptional Activation