Slow activation of fast mitochondrial Ca²⁺ uptake by cytosolic Ca²

Mitochondrial Ca²⁺ uptake through the mitochondrial Ca²⁺ uniporter (MCU) is a tightly controlled process that sustains cell functions mainly by fine-tuning oxidative metabolism to cellular needs. The kinetics of Ca²⁺ fluxes across the mitochondrial membranes have been studied both in vitro and in vivo for many years, and the discovery of the molecular components of the MCU has further clarified that this Ca²⁺ uptake mechanism is based on a complex system subject to elaborate layers of controls. Alterations in the speed or capacity of the in-and-out pathways can have detrimental consequences for both the organelle and the cell, impairing cellular metabolism and ultimately causing cell death. Here, we report that pretreatment of deenergized mitochondria with low-micromolar Ca²⁺ concentrations for a few minutes markedly increases the speed of mitochondrial Ca²⁺ uptake upon re-addition of an oxidizable substrate. We found that this phenomenon is sensitive to alterations in the level of the MCU modulator proteins mitochondrial calcium uptake 1 (MICU1) and 2 (MICU2), and is accompanied by changes in the association of MICU1-MICU2 complexes with MCU. This increased Ca²⁺ uptake capacity, occurring under conditions mimicking those during ischemia/reperfusion in vivo, could lead to a massive amount of Ca²⁺ entering the mitochondrial matrix even at relatively low levels of cytosolic Ca²⁺ We conclude that the phenomenon uncovered here represents a potential threat of mitochondrial Ca²⁺ overload to the cell.