Human immunodeficiency virus type 1 (HIV-1) provirus expression and LTR transcription are repressed in NEF-expressing cell lines.
Human immunodeficiency virus type 1 (HIV-1) NEF protein has been demonstrated to be a negative regulator of HIV-1 replication and HIV-1 LTR transcription under transient expression conditions. The difficulty of several laboratories to reproduce these findings led us to reexamine the role of NEF in HIV-1 provirus expression and HIV-1 LTR transcription. Basal transcription from the HIV-1 LTR in the presence of a NEF expression vector was compared to that in the presence of a mutated NEF vector. NEF expression led to a greater than 10-fold repression of LTR transcription under these conditions. HeLa and Jurkat cell lines carrying the nef gene linked to the CMV promoter or the HIV-1 LTR were isolated by coselection for neomycin resistance. Single cell isolates were further selected for the expression of nef transcripts. With the exception of the anti-sense nef cell lines, all the nef cell lines expressed the 27-kDa NEF protein, detectable by immunoprecipitation. NEF+ HeLa cell lines were at least 5-fold less efficient than NEF- HeLa cell lines in transient proviral expression. Provirus expression was also repressed in the NEF+ Jurkat cell lines. TAT-activated LTR transcription from an HIV-1 LTR-linked CAT expression vector was repressed 10-fold in the NEF+ HeLa and NEF+ Jurkat cell lines. When infected with HIV-1, NEF expressing T lymphoid cell lines showed moderate delays in onset and peak of reverse transcriptase production. However, none of these cell lines completely arrested virus replication. Our data confirm a negative regulatory effect of NEF on both virus production and LTR driven CAT expression in the cell lines tested. It is possible that cell specific factors may influence NEF activity.
Maitra
R K
RK
Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892.
Ahmad
N
N
Holland
S M
SM
Venkatesan
S
S
D000818:Animals; D002460:Cell Line; D015967:Gene Expression Regulation, Viral; D015693:Gene Products, nef / Q000502:physiology*; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D011533:Proviruses / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D051381:Rats; D012097:Repressor Proteins / Q000502:physiology; D013601:T-Lymphocytes / Q000502:physiology; D014158:Transcription, Genetic; D014162:Transfection; D014779:Virus Replication; D054311:nef Gene Products, Human Immunodeficiency Virus
D015693:Gene Products, nef; D012367:RNA, Viral; D012097:Repressor Proteins; D054311:nef Gene Products, Human Immunodeficiency Virus