Journal of Virology, November 1999, p. 9021-9028, Vol. 73, No. 11
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
andFels Institute for Cancer Research and Molecular Biology1 and Department of Biochemistry,2 Temple University School of Medicine, Philadelphia, Pennsylvania 19140
Received 14 April 1999/Accepted 6 August 1999
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-
)-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
subunit of eukaryotic initiation factor 2 and NF-
B 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-
-induced HIV-1 replication.
Present address: Department of Molecular Genetics and Microbiology,
UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854.
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