Mitochondrial division is an important cellular process in both normal and pathological conditions. The dynamin GTPase Drp1 is a central mitochondrial division protein, driving constriction of the outer mitochondrial membrane. In mammals, the outer mitochondrial membrane protein Mff is a key receptor for recruiting Drp1 from the cytosol to the mitochondrion. Actin filaments are also important in Drp1 recruitment and activation. The manner in which Mff and actin work together in Drp1 activation is unknown. Here, we show that Mff is an oligomer (most likely a trimer) that dynamically associates and disassociates through its C-terminal coiled-coil, with a K_d in the range of 10 µM. Dynamic Mff oligomerization is required for Drp1 activation. While not binding Mff directly, actin filaments enhance Mff-mediated Drp1 activation by lowering the effective Mff concentration 10-fold. Total internal reflection microscopy assays using purified proteins show that Mff interacts with Drp1 on actin filaments in a manner dependent on Mff oligomerization. In U2OS cells, oligomerization-defective Mff does not effectively rescue three defects in Mff knock-out cells: mitochondrial division, mitochondrial Drp1 recruitment, and peroxisome division. The ability of Mff to assemble into puncta on mitochondria depends on its oligomerization, as well as on actin filaments and Drp1.

线粒体分裂是正常和病理条件下的重要细胞过程。dynamin GTPase Drp1 是一种中央线粒体分裂蛋白，驱动线粒体外膜的收缩。在哺乳动物中，线粒体外膜蛋白 Mff 是将 Drp1 从胞质溶胶募集到线粒体的关键受体。肌动蛋白丝在 Drp1 募集和激活中也很重要。Mff 和肌动蛋白在 Drp1 激活中协同工作的方式尚不清楚。在这里，我们证明 Mff 是一种寡聚体（很可能是三聚体），它通过其 C 端卷曲线圈动态结合和解离，具有 K _d在 10 µM 范围内。Drp1 激活需要动态 Mff 寡聚化。虽然不直接结合 Mff，但肌动蛋白丝通过将有效 Mff 浓度降低 10 倍来增强 Mff 介导的 Drp1 活化。使用纯化蛋白质的全内反射显微镜分析表明，Mff 与肌动蛋白丝上的 Drp1 以依赖于 Mff 寡聚化的方式相互作用。在 U2OS 细胞中，寡聚化缺陷型 Mff 不能有效挽救 Mff 敲除细胞中的三个缺陷：线粒体分裂、线粒体 Drp1 募集和过氧化物酶体分裂。Mff 在线粒体上组装成斑点的能力取决于它的寡聚化，以及肌动蛋白丝和 Drp1。