Many proteins have been implicated into a multistep process that generates bone-resorbing multinucleated osteoclasts. Here, for the first time, we specifically focus on the cell fusion step of this process. To uncouple merger of the cell membranes from pre-fusion stages of osteoclastogenesis, we accumulated ready-to-fuse cells in the presence of a fusion inhibitor lysophosphatidylcholine and then removed it to study synchronized cell fusion. Synchronized fusion depended on a non-apoptotic exposure of phosphatidylserine at the surface of fusion-committed osteoclast precursors that we found to involve DC-STAMP, a known regulator of osteoclastogenesis. Fusion also depended on cell-surface associated phosphatidylserine-binding proteins annexins (A1 and A5 for murine cells and A5 for human cells). In human osteoclasts, annexin A5 and annexin-binding protein S100A4 regulated fusogenic activity of syncytin 1, another protein that we show to be directly involved in the fusion event. Thus, in contrast to fusion processes mediated by a single protein, such as epithelial cell fusion in C. elegans, cell fusion step in osteoclastogenesis is controlled by a coordinated activity of several proteins.