This proposal will support the candidate's career goals of studying mitochondrial dysfunction caused by abnormal calcium (Ca2+) signaling in cardiac disease. The candidate will use this project to lay the groundwork for this long-term goal by, first, completing necessary experiments to dissect the molecular mechanisms by which Ca2+ uptake occurs, and, second, training in quantitative genomic and bioinformatic methods necessary for isolating patient cohorts possessing such mitochondrial dysfunction. The major mitochondrial protein transporting Ca2+ during signaling events is the mitochondrial Ca2+ uniporter, a channel embedded in the inner membrane. This channel possesses two key features. It is highly selective for Ca2+, not allowing other ions to enter at resting cytoplasmic Ca2+ levels. And it transports Ca2+ only when cytoplasmic levels are high, such as during signaling events or if Ca2+ clearance is insufficient. This selectivity and regulation prevent unnecessary ion transport, which would lead to mitochondrial uncoupling and failure, and they may be altered in heart disease. To identify how the channel performs these two key functions, a mutational analysis of the recently- discovered genes that form the pore (MCU) and accessory subunits (MICU1) of the channel will be conducted. Prior investigations have been hampered by the use of imaging methods that cannot control for secondary factors influencing Ca2+ uptake, leading to contradictory models. The chief innovation of this proposal is the use of mitochondrial electrophysiology, which controls for precisely these secondary factors. In the mentored phase, experiments will test the hypotheses that highly-conserved residues facing the inter-membrane space serve to bind Ca2+ and form a narrow, rigid pore, preventing the transport of other ions. In the independent phase, experiments will test the hypothesis that the MICU1 subunit inhibits transport at resting cytoplasmic Ca2+ levels by driving the channel into a predominantly closed state, releasing this inhibition during Ca2+ elevations, in contrast to more complicated current models. During the independent phase, the candidate will also receive training in genomic approaches to identify patients with cardiac disease suggesting mitochondrial dysfunction. Modeling this dysfunction in cellular or animal systems will be the basis of future grant applications, to examine in detail to what degree aberrant mitochondrial Ca2+ signaling is causative. In this context, the experiments proposed in this application are necessary to understand how mitochondrial Ca2+ uptake is regulated at baseline. The candidate is well-qualified to carry out the short- and long-term goals described above. He has a strong background in ion-channel biology, has spent considerable effort learning mitochondrial electrophysiology, and plans to conduct his training and research in an environment supported by experts in mitochondrial disease, ion-channel biology, and genomic approaches.
Mitochondria are the energy factories of the cell, and often become dysfunctional in heart failure when overloaded with calcium. In this proposal, we will define how the main portal into mitochondria is able to let in calcium normally without causing mitochondrial failure. Understanding this mechanism will be necessary for future studies defining how this activity is subverted in heart failure.
|Kamer, Kimberli J; Sancak, Yasemin; Fomina, Yevgenia et al. (2018) MICU1 imparts the mitochondrial uniporter with the ability to discriminate between Ca2+ and Mn2+. Proc Natl Acad Sci U S A 115:E7960-E7969|
|Abiria, Sunday A; Krapivinsky, Grigory; Sah, Rajan et al. (2017) TRPM7 senses oxidative stress to release Zn2+ from unique intracellular vesicles. Proc Natl Acad Sci U S A 114:E6079-E6088|
|Sommakia, Salah; Houlihan, Patrick R; Deane, Sadiki S et al. (2017) Mitochondrial cardiomyopathies feature increased uptake and diminished efflux of mitochondrial calcium. J Mol Cell Cardiol 113:22-32|
|Fidler, Trevor P; Campbell, Robert A; Funari, Trevor et al. (2017) Deletion of GLUT1 and GLUT3 Reveals Multiple Roles for Glucose Metabolism in Platelet and Megakaryocyte Function. Cell Rep 20:881-894|
|Chaudhuri, Dipayan; Artiga, Daniel J; Abiria, Sunday A et al. (2016) Mitochondrial calcium uniporter regulator 1 (MCUR1) regulates the calcium threshold for the mitochondrial permeability transition. Proc Natl Acad Sci U S A 113:E1872-80|