Author SummaryMammalian reproduction requires successful meiosis, which consists of two strongly asymmetric cell divisions. In meiosis I, movement of the spindle (the subcellular structure that segregates chromosomes during division) toward the oocyte cortex (the outer layer of the egg) is essential for fertility. This process requires that actin filaments assemble in a dynamic mesh, driven by three actin binding proteins, profilin, formin 2, and Spire. To date the molecular mechanisms by which these three proteins cooperate are not known. We now explore this in vitro by a combination of bulk solution and single actin filament assembly assays in the presence of profilin, Spire, and formin 2. Individually, Spire binds to actin filament ends to block their growth, and by itself, formin 2 associates poorly with filament ends, promoting fast processive assembly from the profilin-actin complex. However, when present together, Spire and formin 2 interact with one another (the formin 2 C-terminal binds to the N terminal Spire KIND domain), forming transient complexes at filament ends from which each binds alternately to the filament ends to regulate actin assembly by a ping-pong mechanism. Our in vitro observations are validated by injection studies in mouse oocytes. In oocytes, the additional interaction of Spire and formin 2 with Rab11a-myosin Vb vesicles couples high actin dynamics to vesicle traffic.