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J. Cell Biol.,
Volume 139, Number 3, November 3, 1997 651-664
* Medical Research Council Laboratory for Molecular Cell Biology and Department of Biochemistry, University College
London, London WC1E 6BT, United Kingdom; The chemokine receptor CXCR4 is required,
together with CD4, for entry by some isolates of HIV-1,
particularly those that emerge late in infection. The use
of CXCR4 by these viruses likely has profound effects
on viral host range and correlates with the evolution of
immunodeficiency. Stromal cell-derived factor-1 (SDF-1), the ligand for CXCR4, can inhibit infection by CXCR4-dependent viruses. To understand the mechanism of
this inhibition, we used a monoclonal antibody that is
specific for CXCR4 to analyze the effects of phorbol esters and SDF-1 on surface expression of CXCR4. On
human T cell lines SupT1 and BC7, CXCR4 undergoes
slow constitutive internalization (1.0% of the cell surface pool/min). Addition of phorbol esters increased
this endocytosis rate >6-fold and reduced cell surface
CXCR4 expression by 60 to 90% over 120 min. CXCR4
was internalized through coated pits and coated vesicles and subsequently localized in endosomal compartments from where it could recycle to the cell surface after removal of the phorbol ester. SDF-1 also induced
the rapid down modulation (half time ~5 min) of
CXCR4. Using mink lung epithelial cells expressing CXCR4 and a COOH-terminal deletion mutant of
CXCR4, we found that an intact cytoplasmic COOH-terminal domain was required for both PMA and
ligand-induced CXCR4 endocytosis. However, experiments using inhibitors of protein kinase C indicated
that SDF-1 and phorbol esters trigger down modulation
through different cellular mechanisms.
SDF-1 inhibited HIV-1 infection of mink cells expressing CD4 and CXCR4. The inhibition of infection
was less efficient for CXCR4 lacking the COOH-terminal domain, suggesting at least in part that SDF-1 inhibition of virus infection was mediated through ligand-induced internalization of CXCR4. Significantly, ligand induced internalization of CXCR4 but not CD4, suggesting that CXCR4 and CD4 do not normally physically interact on the cell surface. Together these studies
indicate that endocytosis can regulate the cell-surface
expression of CXCR4 and that SDF-1-mediated down
regulation of cell-surface coreceptor expression contributes to chemokine-mediated inhibition of HIV infection.
Department of Medicine, University of Pennsylvania, Philadelphia,
Pennsylvania 19104; § Laboratory for Molecular Pharmacology, Rigshospitalet 6321, DK-2100 Copenhagen, Denmark; Division
of Molecular Sciences, Glaxo Research Institute, Research Triangle Park, North Carolina 27709; and ¶ Geneva Biomedical
Research Institute, GlaxoWellcome Research and Development SA, 1228-Plan-Les-Ouabes, Geneva, Switzerland
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