Inhibition of replication of reactivated human immunodeficiency virus type 1 (HIV-1) in latently infected U1 cells transduced with an HIV-1 long terminal repeat-driven PKR cDNA construct.
Treatment of human immunodeficiency virus type 1 (HIV-1)-infected individuals with highly active antiretroviral therapy has effectively decreased viral load to undetectable levels. However, efforts to eliminate HIV-1 from these individuals have been unsuccessful, due to the presence of stable, latent viral reservoirs in resting and active CD4(+) T lymphocytes and macrophages. These latent populations have become critical targets in the effort to eradicate HIV-1 from infected individuals. The mechanisms of HIV-1 latency have been studied by using the HIV-1-infected promonocytic cell line U1. The interferon-inducible double-stranded RNA-dependent p68 protein kinase (PKR), a key enzyme in the host-mediated antiviral response, is known to be down-regulated during HIV-1 infection. Therefore, in order to evaluate the role of PKR in the inhibition of replication of reactivated HIV-1 in latently infected U1 cells, we have utilized cDNA constructs containing PKR under the transcriptional control of the HIV-1 long terminal repeat. One PKR-transduced clone, U1/106-4:27, inhibited the tumor necrosis factor alpha (TNF-alpha)-induced replication of HIV-1 by 99% compared to control U1 cells as measured by syncytium formation and HIV-1 p24 antigen enzyme-linked immunosorbent assay. Western blot analysis showed an increase in PKR expression through 96 h postinduction in the U1/106-4:27 clone, concomitant with maximal increases in phosphorylation of the alpha subunit of eukaryotic initiation factor 2 and NF-kappaB activity at 72 h postinduction. These results demonstrate that overexpression of PKR can inhibit the replication of reactivated HIV-1 in latently infected cells and confirm the involvement of PKR in the interferon-associated antiviral pathway against HIV-1 infection. Additionally, treatment of the PKR-transduced U1/106-4:27 clone with the protease inhibitor saquinavir (250 nM) completely inhibited TNF-alpha-induced HIV-1 replication.
Muto
N F
NF
Fels Institute for Cancer Research, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.
Martinand-Mari
C
C
Adelson
M E
ME
Suhadolnik
R J
RJ
D002460:Cell Line; D018076:DNA, Complementary; D015967:Gene Expression Regulation, Viral; D016325:HIV Long Terminal Repeat / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism; D014161:Transduction, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014775:Virus Activation / Q000187:drug effects; D017735:Virus Latency; D014779:Virus Replication; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*
D018076:DNA, Complementary; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D019892:eIF-2 Kinase