California serogroup Gc (G1) glycoprotein is the principal determinant of pH-dependent cell fusion and entry

In the current study, a novel coreceptor-specific cell-cell fusion (CCF) assay system is reported. The system possesses the following features: dual CCR5-dependent and CXCR4-dependent CCF assays, all stable cell lines, inducible expression of gp160 to minimize cytotoxicity, robust luciferase reporter, and 384-well format. These assays have been validated using various known HIV entry inhibitors targeting various stages of the HIV entry/fusion process, including fusion inhibitors, gp120 inhibitors, CCR5 antagonists, CCR5 antibodies, and CXCR4 antagonists. IC 50data generated from this assay system were well correlated to that from the antiviral assays. The effects of DMSOon this assay systemwere assessed, and a 2-to 3-fold increase in luciferase activitywas observed in the presence of 0.05% to2% DMSO. Although cell-cell fusion efficiencywas enhanced, no changes in drug response kinetics for entry inhibitors were found in the presence of 0.1% or 0.5% DMSO. This assay system has been successfully used for the identification and characterization of thousands of CCR5 inhibitors.

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Human immunodeficiency virus envelope-dependent cell-cell fusion: A quantitative fluorescence cytometric assay

Cytometry ◽

10.1002/cyto.10051 ◽

2002 ◽

Vol 47 (2) ◽

pp. 100-106 ◽

Cited By ~ 31

Author(s):

Leonor Huerta ◽

Edmundo Lamoyi ◽

Armida Báez-Saldaña ◽

Carlos Larralde

Keyword(s):

Human Immunodeficiency Virus ◽

Cell Fusion ◽

Virus Envelope ◽

Human Immunodeficiency Virus Envelope ◽

Immunodeficiency Virus ◽

Dependent Cell ◽

Quantitative Fluorescence ◽

Cell Cell

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pH-Dependent Hemolysis and Cell Fusion of Rhabdoviruses

Microbiology and Immunology ◽

10.1111/j.1348-0421.1982.tb00252.x ◽

1982 ◽

Vol 26 (11) ◽

pp. 1035-1043 ◽

Cited By ~ 6

Author(s):

Kazuaki Mannen ◽

Masanobu Ohuchi ◽

Kumato Mifune

Keyword(s):

Cell Fusion ◽

Ph Dependent

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The Avian Coronavirus Infectious Bronchitis Virus Undergoes Direct Low-pH-Dependent Fusion Activation during Entry into Host Cells

Journal of Virology ◽

10.1128/jvi.80.7.3180-3188.2006 ◽

2006 ◽

Vol 80 (7) ◽

pp. 3180-3188 ◽

Cited By ~ 49

Author(s):

Victor C. Chu ◽

Lisa J. McElroy ◽

Vicky Chu ◽

Beverley E. Bauman ◽

Gary R. Whittaker

Keyword(s):

Cell Fusion ◽

Conformational Changes ◽

Infectious Bronchitis Virus ◽

Fusion Reaction ◽

Low Ph ◽

Surface Proteins ◽

Host Cells ◽

Dependent Manner ◽

Infectious Bronchitis ◽

Ph Dependent

ABSTRACT Coronaviruses are the causative agents of respiratory disease in humans and animals, including severe acute respiratory syndrome. Fusion of coronaviruses is generally thought to occur at neutral pH, although there is also evidence for a role of acidic endosomes during entry of a variety of coronaviruses. Therefore, the molecular basis of coronavirus fusion during entry into host cells remains incompletely defined. Here, we examined coronavirus-cell fusion and entry employing the avian coronavirus infectious bronchitis virus (IBV). Virus entry into cells was inhibited by acidotropic bases and by other inhibitors of pH-dependent endocytosis. We carried out fluorescence-dequenching fusion assays of R18-labeled virions and show that for IBV, coronavirus-cell fusion occurs in a low-pH-dependent manner, with a half-maximal rate of fusion occurring at pH 5.5. Fusion was reduced, but still occurred, at lower temperatures (20°C). We observed no effect of inhibitors of endosomal proteases on the fusion event. These data are the first direct measure of virus-cell fusion for any coronavirus and demonstrate that the coronavirus IBV employs a direct, low-pH-dependent virus-cell fusion activation reaction. We further show that IBV was not inactivated, and fusion was unaffected, by prior exposure to pH 5.0 buffer. Virions also showed evidence of reversible conformational changes in their surface proteins, indicating that aspects of the fusion reaction may be reversible in nature.

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