The Mechanism of MICU-Dependent Gating of the Mitochondrial Ca²⁺Uniporter

Mitochondrial Ca²⁺uniporter (MCU) mediates mitochondrial Ca²⁺uptake, regulating ATP production and cell death. According to the existing paradigm, MCU is occluded at the resting cytosolic [Ca²⁺] and only opens above an ∼400 nM threshold. This Ca²⁺-dependent gating is putatively conferred by MICUs, EF hand-containing auxiliary subunits that block/unblock the MCU pore depending on cytosolic [Ca²⁺]. Here we provide the first direct, patch-clamp based analysis of the Ca²⁺-dependent MCU gating and the role played by MICUs. Surprisingly, MICUs do not occlude the MCU pore, and MCU is a constitutively active channel without cytosolic [Ca²⁺] activation threshold. Instead, MICUs potentiate MCU activity when cytosolic Ca²⁺binds to their EF hands. MICUs cause this potentiation by increasing the probability of open state of the MCU channel.