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Quantitative 3D video microscopy of HIV transfer across T cell virological synapses.

ABSTRACT: The spread of HIV between immune cells is greatly enhanced by cell-cell adhesions called virological synapses, although the underlying mechanisms have been unclear. With use of an infectious, fluorescent clone of HIV, we tracked the movement of Gag in live CD4 T cells and captured the direct translocation of HIV across the virological synapse. Quantitative, high-speed three-dimensional (3D) video microscopy revealed the rapid formation of micrometer-sized "buttons" containing oligomerized viral Gag protein. Electron microscopy showed that these buttons were packed with budding viral crescents. Viral transfer events were observed to form virus-laden internal compartments within target cells. Continuous time-lapse monitoring showed preferential infection through synapses. Thus, HIV dissemination may be enhanced by virological synapse-mediated cell adhesion coupled to viral endocytosis.

SUBMITTER: Hubner W

PROVIDER: S-EPMC2756521 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Quantitative 3D video microscopy of HIV transfer across T cell virological synapses.

Hübner Wolfgang W McNerney Gregory P GP Chen Ping P Dale Benjamin M BM Gordon Ronald E RE Chuang Frank Y S FY Li Xiao-Dong XD Asmuth David M DM Huser Thomas T Chen Benjamin K BK

Science (New York, N.Y.) 20090301 5922

The spread of HIV between immune cells is greatly enhanced by cell-cell adhesions called virological synapses, although the underlying mechanisms have been unclear. With use of an infectious, fluorescent clone of HIV, we tracked the movement of Gag in live CD4 T cells and captured the direct translocation of HIV across the virological synapse. Quantitative, high-speed three-dimensional (3D) video microscopy revealed the rapid formation of micrometer-sized "buttons" containing oligomerized viral ...[more]

PMID: 19325119

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Project description:UnlabelledRetroviruses spread more efficiently when infected and uninfected cells form tight, physical interfaces known as virological synapses (VSs). VS formation is initiated by adhesive interactions between viral Envelope (Env) glycoproteins on the infected cell and CD4 receptor molecules on the uninfected cell. How high-avidity Env-CD4 linkages are resolved over time is unknown. We describe here a tractable two-color, long-term (>24 h) live cell imaging strategy to study VS turnover in the context of a large cell population, quantitatively. We show that Env's conserved cytoplasmic tail (CT) can potently signal the recruitment of Gag capsid proteins to the VS, a process also dependent on residues within Gag's N-terminal matrix (MA) domain. Additionally, we demonstrate that Env's CT and Gag's MA domain both regulate the duration of interactions between viral donor and target cells, as well as the stability of this interaction over time (i.e., its capacity to resolve or form a syncytium). Finally, we report the unexpected finding that modulating extracellular fluid viscosity markedly impacts target T cell trafficking and thus affects the duration, stability, and turnover of virus-induced cell-cell contacts. Combined, these results suggest a stepwise model for viral cell-to-cell transmission wherein (i) Env-receptor interactions anchor target cells to infected cells, (ii) Env signals Gag's recruitment to the cell-cell contact dependent on an intact Env CT and Gag MA, and (iii) Env CT and Gag MA, in conjunction with extracellular forces, combine to regulate VS stability and infectious outcomes.ImportanceHIV-1 spreads efficiently at physical, cell-cell interfaces known as virological synapses (VSs). The VS provides for spatiotemporal coupling of virus assembly and entry into new host cells and may transmit signals relevant to pathogenesis. Disrupting this mode of transmission may be critical to the goal of abolishing viral persistence in infected individuals. We describe here a long-term live cell imaging strategy for studying virus-induced effects on cell behavior in the context of a large cell population. We demonstrate cooperative roles for viral Gag capsid proteins and Envelope glycoproteins in regulating VS formation and turnover. We also show that modulating fluid viscosity markedly affects T cell trafficking and VS stability. Thus, extracellular factors also play an important role in modulating the nature of infectious cell-cell interactions. In sum, our study provides new tools and insights relevant to exposing vulnerabilities in how HIV-1 and other viruses spread infection among cells, tissues, and people.

Dataset Information

Quantitative 3D video microscopy of HIV transfer across T cell virological synapses.

Publications

Quantitative 3D video microscopy of HIV transfer across T cell virological synapses.

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