Identification of Critical MCUR1 Domains in the Mitochondrial Calcium Uniporter Complex that Regulates Cellular Metabolism

Abstract

Mitochondria are multivariate signal processors that decode diverse cellular inputs into various bioenergetic outputs. One such signal is the calcium (Ca2+) flux between the cytosol and mitochondria which modulates the organelle’s function. Under physiologic cytosolic [Ca2+], energy output (ATP) can be adjusted to meet cellular demands. To maintain the Ca2+ homeostatic circuit, a selective mitochondrial Ca2+ Uniporter (MCU) complex promotes Ca2+ uptake. MCUR1 is an integral mitochondrial inner membrane protein which serves as a scaffold factor for the MCU complex and regulates its functional activity. The resolved NMR structure of the human MCUR1 protein revealed a high degree of conservation for 3 residues within the “head domain” of the protein, A170, L174, V203, and Y204.ObjectiveWe hypothesized that alterations of these critical residues would affect MCUR1 function, and thereby, MCU complex activity.MethodsTo test this hypothesis, MCU‐mediated mitochondrial calcium (mCa2+) uptake, resting matrix mCa2+ levels, and mitochondrial bioenergetics such as NAD(P)H production and oxygen consumption rate (OCR) were evaluated.ResultsRemarkably, cells expressing the MCUR1A170R/L174R mutant demonstrated near complete ablation of MCU‐mediated mCa2+ uptake, NAD(P)H production and OCR. Conversely, cells expressing MCURWT, or the MCURV203R/Y204R mutant demonstrated similar effects on MCU activity and mitochondrial bioenergetics suggesting a nominal change at the structural level.ConclusionCollectively, our results indicate that the MCUR1 head domain is necessary for proper MCU complex activity, and that MCU‐mediated mCa2+ uptake modulates mitochondrial bioenergetics and cell function.Support or Funding InformationThis research was funded by the National Institutes of Health R25R25DK078381 program. (NIHR01GM109882, R01HL086699, R01HL142673, DOD/CDMRP PR181598P and NIH1S10RR027327 to M. Madesh and R25DK078381 to W.B.R. B.T.E. is supported by the National Center For Advancing Translational Sciences of the National Institutes of Health under Award Number TL1TR002647. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.