Ferroptosis triggers mitochondrial fragmentation via Drp1 activation

Constitutive mitochondrial dynamics ensure quality control and metabolic fitness of cells, and their dysregulation has been implicated in various human diseases. The large GTPase Dynamin-related protein 1 (Drp1) is intimately involved in mediating constitutive mitochondrial fission and has been implicated in mitochondrial cell death pathways. During ferroptosis, a recently identified type of regulated necrosis driven by excessive lipid peroxidation, mitochondrial fragmentation has been observed. Yet, how this is regulated and whether it is involved in ferroptotic cell death has remained unexplored. Here, we provide evidence that Drp1 is activated upon experimental induction of ferroptosis and promotes cell death execution and mitochondrial fragmentation. Using time-lapse microscopy, we found that ferroptosis induced mitochondrial fragmentation and loss of mitochondrial membrane potential, but not mitochondrial outer membrane permeabilization. Importantly, Drp1, the major GTPase involved in steady-state mitochondrial fission, accelerated ferroptotic cell death kinetics. Notably, this function was mediated by regulation of mitochondrial dynamics as overexpression of Mitofusin 2 phenocopied the effect of Drp1 deficiency in delaying ferroptosis cell death kinetics. Mechanistically, we found that Drp1 is phosphorylated and activated after induction of ferroptosis and translocated to mitochondria. Importantly, recruitment to and its further activation at mitochondria through the phosphatase PGAM5 promoted ferroptotic cell death. These data provide the first evidence for a functional role of Drp1 activation in the acceleration of ferroptotic cell death and mitochondrial fragmentation, with important implications for the regulation of mitochondrial dynamics in diseases associated with ferroptosis.