Quantitative multicolor super-resolution microscopy reveals tetherin HIV-1 interaction

• Published in: PLoS pathogens Vol. 7; no. 12; p. e1002456
• Main Authors: Lehmann, Martin, Rocha, Susana, Mangeat, Bastien, Blanchet, Fabien, Uji-I, Hiroshi, Hofkens, Johan, Piguet, Vincent
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 01.12.2011; Public Library of Science (PLoS)
• Subjects: Adaptive immunology; Antigens, CD - chemistry; Antigens, CD - metabolism; Bars; Biology; Care and treatment; Cell Line; Cell Membrane - immunology; Cell Membrane - metabolism; Cellular Biology; Cluster analysis; Color; Fluorescent Antibody Technique - methods; Gangliosides; Genetic aspects; GPI-Linked Proteins - chemistry; GPI-Linked Proteins - metabolism; HIV; HIV infection; HIV-1 - metabolism; Human immunodeficiency virus; Human Immunodeficiency Virus Proteins - metabolism; Humans; Immunology; Innate immunity; Labeling; Life Sciences; Light; Lipids; Microbiology and Parasitology; Microscope and microscopy; Microscopy; Microscopy, Confocal; Microscopy, Electron, Transmission - methods; Molecular weight; Physiological aspects; Plasma; Proteins; Transfection; Virion - immunology; Virion - metabolism; Virology; Viruses; United Kingdom; Switzerland
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1002456

Virus assembly and interaction with host-cell proteins occur at length scales below the diffraction limit of visible light. Novel super-resolution microscopy techniques achieve nanometer resolution of fluorescently labeled molecules. The cellular restriction factor tetherin (also known as CD317, BST-2 or HM1.24) inhibits the release of human immunodeficiency virus 1 (HIV-1) through direct incorporation into viral membranes and is counteracted by the HIV-1 protein Vpu. For super-resolution analysis of HIV-1 and tetherin interactions, we established fluorescence labeling of HIV-1 proteins and tetherin that preserved HIV-1 particle formation and Vpu-dependent restriction, respectively. Multicolor super-resolution microscopy revealed important structural features of individual HIV-1 virions, virus assembly sites and their interaction with tetherin at the plasma membrane. Tetherin localization to micro-domains was dependent on both tetherin membrane anchors. Tetherin clusters containing on average 4 to 7 tetherin dimers were visualized at HIV-1 assembly sites. Combined biochemical and super-resolution analysis revealed that extended tetherin dimers incorporate both N-termini into assembling virus particles and restrict HIV-1 release. Neither tetherin domains nor HIV-1 assembly sites showed enrichment of the raft marker GM1. Together, our super-resolution microscopy analysis of HIV-1 interactions with tetherin provides new insights into the mechanism of tetherin-mediated HIV-1 restriction and paves the way for future studies of virus-host interactions.