Late stages of the synchronized macrophage fusion in osteoclast formation depend on dynamin

Macrophage fusion that leads to osteoclast formation is one of the most important examples of cell-to-cell fusion in development, tissue homeostasis and immune response. Protein machinery that fuses macrophages remains to be identified. Here we explored fusion stage of osteoclast formation for RAW macrophage-like murine cells and for macrophages derived from human monocytes. To uncouple fusion from the preceding differentiation processes, we accumulated fusion-committed cells in the presence of lysophosphatidylcholine (LPC) that reversibly blocks membrane merger. 16h later we removed LPC and observed cell fusion events that would normally develop within 16h to develop within 30-90 min. Thus, while osteoclastogenesis, generally, takes several days, our approach allowed us to focus on an hour, in which we observe robust fusion between the cells. Complementing syncytium formation assay with a novel membrane merger assay let us study the synchronized fusion events downstream of a local merger between two plasma membranes but before expansion of nascent membrane connections and complete unification of the cells. We found that the expansion of membrane connections detected as a growth of multinucleated osteoclasts depends on dynamin activity. In contrast, a merger between the plasma membranes of the two cells was not affected by inhibitors of dynamin GTPase. Thus, dynamin that was recently found to control late stages of myoblast fusion also controls late stages of macrophage fusio...
Source: BJ Cell - Category: Biochemistry Authors: Tags: BJ Cell Source Type: research