MICU1 controls both the threshold and cooperative activation of the mitochondrial Ca²⁺ uniporter.

Mitochondrial Ca(2+) uptake via the uniporter is central to cell metabolism, signaling, and survival. Recent studies identified MCU as the uniporter's likely pore and MICU1, an EF-hand protein, as its critical regulator. How this complex decodes dynamic cytoplasmic [Ca(2+)] ([Ca(2+)]c) signals, to tune out small [Ca(2+)]c increases yet permit pulse transmission, remains unknown. We report that loss of MICU1 in mouse liver and cultured cells causes mitochondrial Ca(2+) accumulation during small [Ca(2+)]c elevations but an attenuated response to agonist-induced [Ca(2+)]c pulses. The latter reflects loss of positive cooperativity, likely via the EF-hands. MICU1 faces the intermembrane space and responds to [Ca(2+)]c changes. Prolonged MICU1 loss leads to an adaptive increase in matrix Ca(2+) binding, yet cells show impaired oxidative metabolism and sensitization to Ca(2+) overload. Collectively, the data indicate that MICU1 senses the [Ca(2+)]c to establish the uniporter's threshold and gain, thereby allowing mitochondria to properly decode different inputs.