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

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. © 2011 lehmann et al.