The MCU and MCUb amino-terminal domains tightly interact: mechanisms for low conductance assembly of the mitochondrial calcium uniporter complex

Megan Noble; Danielle M Colussi; Murray Junop; Peter B Stathopulos

doi:10.1016/j.isci.2024.109699

The MCU and MCUb amino-terminal domains tightly interact: mechanisms for low conductance assembly of the mitochondrial calcium uniporter complex

iScience. 2024 Apr 10;27(5):109699. doi: 10.1016/j.isci.2024.109699. eCollection 2024 May 17.

Authors

Megan Noble¹, Danielle M Colussi¹, Murray Junop², Peter B Stathopulos¹

Affiliations

¹ Department of Physiology and Pharmacology, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON N6A5C1, Canada.
² Department of Biochemistry, Schulich School of Medicine and Dentistry, University of Western Ontario, London, ON N6A5C1, Canada.

Abstract

The mitochondrial calcium (Ca²⁺) uniporter (MCU) complex is regulated via integration of the MCU dominant negative beta subunit (MCUb), a low conductance paralog of the main MCU pore forming protein. The MCU amino (N)-terminal domain (NTD) also modulates channel function through cation binding to the MCU regulating acidic patch (MRAP). MCU and MCUb have high sequence similarities, yet the structural and functional roles of MCUb-NTD remain unknown. Here, we report that MCUb-NTD exhibits α-helix/β-sheet structure with a high thermal stability, dependent on protein concentration. Remarkably, MCU- and MCUb-NTDs heteromerically interact with ∼nM affinity, increasing secondary structure and stability and structurally perturbing MRAP. Further, we demonstrate MCU and MCUb co-localization is suppressed upon NTD deletion concomitant with increased mitochondrial Ca²⁺ uptake. Collectively, our data show that MCU:MCUb NTD tight interactions are promoted by enhanced regular structure and stability, augmenting MCU:MCUb co-localization, lowering mitochondrial Ca²⁺ uptake and implicating an MRAP-sensing mechanism.

Keywords: Cell biology; Structural biology.