TY - JOUR T1 - EWI-2 Inhibits Cell-Cell Fusion at the HIV-1 Virological Presynapse JF - bioRxiv DO - 10.1101/296251 SP - 296251 AU - Emily E. Whitaker AU - Nicholas J. Matheson AU - Sarah Perlee AU - Phillip B. Munson AU - Menelaos Symeonides AU - Markus Thali Y1 - 2019/01/01 UR - http://biorxiv.org/content/early/2019/10/15/296251.abstract N2 - Cell-to-cell transfer of virus particles at the Env-dependent virological synapse (VS) is a highly efficient mode of HIV-1 transmission. While cell-cell fusion could be triggered at the VS, leading to the formation of syncytia and preventing exponential growth of the infected cell population, this is strongly inhibited by both viral (Gag) and host (ezrin and tetraspanins) proteins. Here, we identify EWI-2, a protein that was previously shown to associate with ezrin and tetraspanins, as a host factor that contributes to the inhibition of Env-mediated cell-cell fusion. Using quantitative fluorescence microscopy, shRNA knockdowns, and cell-cell fusion assays, we show that EWI-2 accumulates at the presynaptic terminal (i.e. the producer cell side of the VS), where it contributes to the fusion-preventing activities of the other viral and cellular components. We also find that EWI-2, like tetraspanins, is downregulated upon HIV-1 infection, mostly by Vpu. Despite strong inhibition of fusion at the VS, T cell-based syncytia do form in vivo and in physiologically relevant culture systems, but they remain small. In regard to that, we demonstrate that EWI-2 and CD81 levels are restored on the surface of syncytia, where they (presumably) continue to act as fusion inhibitors. This study documents a new role for EWI-2 as an inhibitor of HIV-1-induced cell-cell fusion, and provides novel insight into how syncytia are prevented from fusing indefinitely. ER -