MICU1 occludes MCU in the mitochondrial calcium uniporter complex

The mitochondrial calcium uniporter imports cytoplasmic Ca2+ into the mitochondrial matrix to regulate cell bioenergetics, Ca2+ signaling, and apoptosis. The uniporter contains the pore-forming MCU subunit, an EMRE protein that binds to MCU, and the regulatory MICU1/MICU2 subunits. Structural and biochemical studies have suggested that MICU1 gates MCU by blocking and unblocking the Ca2+ pore. However, mitoplast patch-clamp experiments argue that MICU1 does not block Ca2+ transport but instead potentiates MCU. To address this direct clash of proposed MICU1 function, we applied purified MICU1 to Ca2+-conducting MCU-EMRE subcomplexes in outside-out patches excised from Xenopus oocytes. MICU1 strongly inhibits Ca2+ currents, and the inhibition is abolished by mutating an MCU-interacting K126 residue in MICU1. Further experiments show that MICU1 block was not observed in mitoplasts because MICU1 dissociates from the uniporter complex. These results firmly establish that MICU1 shuts the uniporter in resting cellular conditions.