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Journal of Virology, March 2001, p. 2993-3000, Vol. 75, No. 6
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.6.2993-3000.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Human Immunodeficiency Virus Type 1 Nef Functions at
the Level of Virus Entry by Enhancing Cytoplasmic Delivery of
Virions
Evelyne
Schaeffer,1,2
Romas
Geleziunas,1 and
Warner C.
Greene1,3,*
Gladstone Institute of Virology and
Immunology1 and Departments of Medicine
and of Microbiology and Immunology,3
University of California, San Francisco, California, and
Unité 338 INSERM, 67084 Strasbourg Cedex,
France2
Received 11 October 2000/Accepted 7 December 2000
The Nef protein of the type 1 human immunodeficiency
virus (HIV-1) plays a key although poorly understood role in
accelerating the progression of clinical disease in vivo. Nef exerts
several biological effects in vitro, including enhancement of
virion infectivity, downregulation of CD4 and major histocompatibility
complex class I receptor expression, and modulation of various
intracellular signaling pathways. The positive effect of Nef on virion
infectivity requires its expression in the producer cell, although its
effect is manifested in the subsequent target cell of infection. Prior studies suggest that Nef does not alter viral entry into target cells;
nevertheless, it enhances proviral DNA synthesis, arguing for an action
of Nef at the level of viral uncoating or reverse transcription. However, these early studies discounting an effect of Nef on virion entry may be confounded by the
recent finding that HIV enters cells by both fusion and
endocytosis. Using epifluorescence microscopy to monitor green
fluorescent protein-Vpr-labeled HIV virion entry into HeLa cells, we
find that endocytosis forms a very active pathway for virus uptake.
Virions entering via the endocytic pathway do not support productive
infection of the host cell, presumably reflecting their inability to
escape from the endosomes. Conversely, our studies now demonstrate that
HIV Nef significantly enhances CD4- and chemokine receptor-dependent
entry of HIV virions into the cytoplasmic compartment of target
cells. Mutations in Nef either impairing its ability to downregulate CD4 or disrupting its polyproline helix compromise virion entry into
the cytoplasm. We conclude that Nef acts at least in part as a
regulator of cytosolic viral entry and that this action contributes to
its positive effects on viral infectivity.
*
Corresponding author. Mailing address: Gladstone
Institute of Virology and Immunology, P.O. Box 419100, San
Francisco, CA 94141-9100. Phone: (415) 695-3800. Fax: (415)
826-1817. E-mail: wgreene{at}gladstone.ucsf.edu.
Journal of Virology, March 2001, p. 2993-3000, Vol. 75, No. 6
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.6.2993-3000.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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