Mitochondrial calcium (Ca2+) signaling has long been known to regulate diverse cellular functions, ranging from ATP production via oxidative phosphorylation, to cytoplasmic Ca2+signaling to apoptosis. Central to mitochondrial Ca2+signaling is the mitochondrial Ca2+uniporter complex (MCUC) which enables Ca2+flux from the cytosol into the mitochondrial matrix. Several pivotal discoveries over the past 15 years have clarified the identity of the proteins comprising MCUC. Here, we provide an overview of the literature on mitochondrial Ca2+biology and highlight recent findings on the high-resolution structure, dynamic regulation, and new functions of MCUC, with an emphasis on publications from the last five years. We discuss the importance of these findings for human health and the therapeutic potential of targeting mitochondrial Ca2+signaling.