Medizinische Universität Graz - Research portal
Selected Publication:
Parichatikanond, W.
Studying the complexity of mitochondrial Ca2+ uptake: Structural reorganization of mitochondrial Ca2+ uptake 1 (MICU1) multimers for activation of mitochondrial Ca2+ uniporter (MCU) is exclusively controlled by cytosolic Ca2+
Doktoratsstudium der Medizinischen Wissenschaft; Humanmedizin; [ Dissertation ] Graz Medical University; 2016. pp.
[OPEN ACCESS]
FullText
- Authors Med Uni Graz:
- Advisor:
- Graier Wolfgang
- Groschner Klaus
- Malli Roland
- Altmetrics:
- Abstract:
- Mitochondrial Ca2+ uptake plays an important role in the regulation of cellular signaling, aerobic metabolism, and apoptosis. Mitochondria are able to buffer Ca2+ from locally high cytosolic Ca2+ levels that modulates various Ca2+ signaling events. Recently, several proteins located in the inner mitochondrial membrane (IMM) have been described to be essential for mitochondrial Ca2+ machineries, including mitochondrial Ca2+ uniporter (MCU), mitochondrial Ca2+ uptake 1 (MICU1), uncoupling proteins 2 and 3 (UCP2/3), essential MCU regulator (EMRE), and mitochondrial Ca2+ uniporter regulator 1 (MCUR1). Although the characteristics and physiological consequences of mitochondrial Ca2+ sequestration have been reported, the actual mechanisms of protein-protein interactions involved in this phenomenon are still unclear. In this study, overexpression and/or silencing of the distinct proteins were applied to reveal that mitochondrial Ca2+ uptake upon IP3-mediated depletion of the endoplasmic reticulum (ER) Ca2+ stores works exclusively via MCU and EMRE but not MCUR1. MICU1 functions as negative regulator of the MCU/EMRE complex. In addition, UCP2/3 facilitates MCU/EMRE functions and counteracts MICU1, while diminution of MICU1 boosts the effect of UCP2/3 on MCU activity in MCU knockdown cells overexpressed with UCP2/3. Our results indicate that UCP2/3 facilitates mitochondrial Ca2+ uptake by MCU/EMRE and thereby controls the mitochondrial Ca2+ entry by function of MICU1. The crystal structure of MICU1 has been recently revealed that in the absence of Ca2+ MICU1 exists as a hexamer, while Ca2+ binding to the two EF-hands results in a rearrangement to MICU1 dimers. Although, the Ca2+-sensing gatekeeper function of MICU1 has been well investigated for MCU that together with the EMRE forms the mitochondrial Ca2+ channel, mechanisms by which MICU1 controls MCU/EMRE activity to tune mitochondrial Ca2+ signals remain ambiguous. Furthermore, overexpression of MICU1 causes pronounced structural alteration of mitochondria pointing to an additional engagement of this protein in the ultrastructure of mitochondria. In this study, we established a Förster-Resonance-Energy-Transfer (FRET)-based live-cell imaging approach to dynamically monitor the kinetics of the structural reorganization of MICU1 oligomers and demonstrate that elevations of cytosolic Ca2+ rearranges MICU1 multimers with an EC50 of approximately 4.4 µM, resulting in activation of mitochondrial Ca2+ uptake via MCU. Moreover, MICU1 rearrangement essentially requires the EF-hand motifs and strictly correlates with the shape of cytosolic Ca2+ rises. Hence, these data show that rearrangements of MICU1 multimers were independent of matrix Ca2+ concentration, mitochondrial membrane potential (¿mito), and expression levels of MCU and EMRE. These data provide novel insights in the dynamics, regulation, and molecular effect of the structural reorganization of MICU1 that adds to the current understanding of the complex molecular mechanisms of MCU/EMRE activation in intact cells.