TMBIM5 is the Ca 2+ /H + antiporter of mammalian mitochondria.

Mitochondrial Ca 2+ ions are crucial regulators of bioenergetics and cell death pathways. Mitochondrial Ca 2+ content and cytosolic Ca 2+ homeostasis strictly depend on Ca 2+ transporters. In recent decades, the major players responsible for mitochondrial Ca 2+ uptake and release have been identified, except the mitochondrial Ca 2+ /H + exchanger (CHE). Originally identified as the mitochondrial K + /H + exchanger, LETM1 was also considered as a candidate for the mitochondrial CHE. Defining the mitochondrial interactome of LETM1, we identify TMBIM5/MICS1, the only mitochondrial member of the TMBIM family, and validate the physical interaction of TMBIM5 and LETM1. Cell-based and cell-free biochemical assays demonstrate the absence or greatly reduced Na + -independent mitochondrial Ca 2+ release in TMBIM5 knockout or pH-sensing site mutants, respectively, and pH-dependent Ca 2+ transport by recombinant TMBIM5. Taken together, we demonstrate that TMBIM5, but not LETM1, is the long-sought mitochondrial CHE, involved in setting and regulating the mitochondrial proton gradient. This finding provides the final piece of the puzzle of mitochondrial Ca 2+ transporters and opens the door to exploring its importance in health and disease, and to developing drugs modulating Ca 2+ exchange.