HIV-Cell Membrane Fusion Intermediates are Restricted by Serinc3 and Serinc5

Abstract

Serinc3 and Serinc5 are recently described host restriction factors that can block HIV infection by incorporating into budding viral particles and decreasing their ability to infect subsequent cells. Serincs are thought to block the very earliest stages of infection, membrane fusion and cell entry, by an incompletely understood mechanism. Understanding how Serincs disrupt HIV membrane fusion will clarify the requirements for normal HIV membrane fusion and could identify new viral weaknesses and drug targets. We used giant plasma membrane vesicles (blebs) as target membranes to study “wildtype” and Serinc-disrupted HIV membrane fusion at a single-particle level by fluorescence microscopy and cryo-Electron Tomography (cryoET). Using fluorescent reporters of membrane and content mixing, we observed that Serinc3 and Serinc5 cause no defect in mixing of the outer lipid leaflets (hemifusion), but a pronounced defect in fusion pore opening. Additionally, cryo-ET of HIV pseudoviruses mixed with blebs allowed observation of multiple intermediates of HIV membrane fusion. We found that Serinc3 and Serinc5 increased the number of hemifusion and early fusion product events observed in these tomograms and many of the fusion products have a narrowed neck between former virus and bleb. These results suggest that Serinc3 and Serinc5 restrict HIV membrane fusion, not by inhibiting a particular intermediate step, but by causing broad changes to the energetics of fusion that enables more frequent observation of multiple transient intermediate states.