Characterization of Mitochondrial Calcium Uniporter in Barth Syndrome Models

Abstract

The Barth Syndrome (BTHS) is caused by the loss of the mitochondrial acyltransferase tafazzin, which is involved in the remodeling process of the hallmark mitochondrial glycerophospholipid cardiolipin. Cardiolipin maintains the stability of various mitochondrial protein complexes, such as respiratory chain complexes and translocases. Our analyses indicate that cardiolipin remodeling plays a crucial role in mitochondrial calcium homeostasis via the Mitochondrial Calcium Uniporter (MCU). MCU is both, a calcium-sensing and a calcium-conducting channel mediating mitochondrial calcium uptake. The mammalian MCU complex consists of the pore-forming subunit MCU, the regulatory constituents MICU1, MICU2 and EMRE, which enable the transition of calcium sensing via MICU1/2 and calcium transduction across the MCU-EMRE-containing pore in the inner mitochondrial membrane. Our current work demonstrates that the assembly and stability of MCU complexes of various molecular weight forms depend on tafazzin function. Using genetic ablation of tafazzin in TAZ KO MEF cells and organs from the BTHS mouse model, we identified three different EMRE-containing MCU oligomers in MEF cells, which undergo a complex ratio shift from WT to TAZ KO MEF mitochondria. Only one of the MCU complexes contains MICU1, which disassembles in TAZ KO MEF cells. This observation correlates with increased basal mitochondrial calcium uptake in TAZ KO MEF cells. Moreover, MCU complexes in BTHS mouse skeletal muscle and brain display similar alterations in complex organization as well. However, MCU complexes are not affected in BTHS mouse hearts, whereas MCU protein levels are reduced. The loss of MCU was recapitulated in BTHS patient-derived cardiomyocytes. Protein steady state levels of MCU in TAZ KO MEF cells are unaffected, whereas the in-vitro import of MCU into purified mitochondria of TAZ KO MEF cells is diminished. Our data display that a reduction of mitochondrial membrane potential affects the in-vitro import of MCU, indicating that MCU precursor import is more sensitive to membrane potential in comparison to control substrates EMRE and OTC. We conclude from our data that the lack of tafazzin function affects the structural integrity of MCU complexes and mitochondrial calcium metabolism.