Letter | Published:

Architecture of the mitochondrial calcium uniporter

Nature volume 533, pages 269273 (12 May 2016) | Download Citation

Abstract

Mitochondria from many eukaryotic clades take up large amounts of calcium (Ca2+) via an inner membrane transporter called the uniporter. Transport by the uniporter is membrane potential dependent and sensitive to ruthenium red or its derivative Ru360 (ref. 1). Electrophysiological studies have shown that the uniporter is an ion channel with remarkably high conductance and selectivity2. Ca2+ entry into mitochondria is also known to activate the tricarboxylic acid cycle and seems to be crucial for matching the production of ATP in mitochondria with its cytosolic demand3. Mitochondrial calcium uniporter (MCU) is the pore-forming and Ca2+-conducting subunit of the uniporter holocomplex, but its primary sequence does not resemble any calcium channel studied to date. Here we report the structure of the pore domain of MCU from Caenorhabditis elegans, determined using nuclear magnetic resonance (NMR) and electron microscopy (EM). MCU is a homo-oligomer in which the second transmembrane helix forms a hydrophilic pore across the membrane. The channel assembly represents a new solution of ion channel architecture, and is stabilized by a coiled-coil motif protruding into the mitochondrial matrix. The critical DXXE motif forms the pore entrance, which features two carboxylate rings; based on the ring dimensions and functional mutagenesis, these rings appear to form the selectivity filter. To our knowledge, this is one of the largest membrane protein structures characterized by NMR, and provides a structural blueprint for understanding the function of this channel.

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Accessions

Primary accessions

Biological Magnetic Resonance Data Bank

Protein Data Bank

Data deposits

The atomic structure coordinate and structural constraints are deposited in the Protein Data Bank (PDB) under the accession number 5ID3. The chemical shift values are deposited in the Biological Magnetic Resonance Data Bank (BMRB) under the accession number 30021.

References

  1. 1.

    & Mechanisms by which mitochondria transport calcium. Am. J. Physiol. 258, C755–C786 (1990)

  2. 2.

    , & The mitochondrial calcium uniporter is a highly selective ion channel. Nature 427, 360–364 (2004)

  3. 3.

    & The role of calcium in the regulation of mitochondrial metabolism. Biochem. Soc. Trans. 8, 266–268 (1980)

  4. 4.

    et al. Integrative genomics identifies MCU as an essential component of the mitochondrial calcium uniporter. Nature 476, 341–345 (2011)

  5. 5.

    et al. MICU1 encodes a mitochondrial EF hand protein required for Ca2+ uptake. Nature 467, 291–296 (2010)

  6. 6.

    , , , & A forty-kilodalton protein of the inner membrane is the mitochondrial calcium uniporter. Nature 476, 336–340 (2011)

  7. 7.

    et al. EMRE is an essential component of the mitochondrial calcium uniporter complex. Science 342, 1379–1382 (2013)

  8. 8.

    et al. The mitochondrial calcium uniporter is a multimer that can include a dominant-negative pore-forming subunit. EMBO J. 32, 2362–2376 (2013)

  9. 9.

    & The molecular era of the mitochondrial calcium uniporter. Nature Rev. Mol. Cell Biol. 16, 545–553 (2015)

  10. 10.

    , , & MCU encodes the pore conducting mitochondrial calcium currents. eLife 2, e00704 (2013)

  11. 11.

    et al. Reconstitution of the mitochondrial calcium uniporter in yeast. Proc. Natl Acad. Sci. USA 111, 8985–8990 (2014)

  12. 12.

    et al. Structure and function of the N-terminal domain of the human mitochondrial calcium uniporter. EMBO Rep. (2015)

  13. 13.

    et al. Solution nuclear magnetic resonance structure of membrane-integral diacylglycerol kinase. Science 324, 1726–1729 (2009)

  14. 14.

    & Structure and mechanism of the M2 proton channel of influenza A virus. Nature 451, 591–595 (2008)

  15. 15.

    et al. Unusual architecture of the p7 channel from hepatitis C virus. Nature 498, 521–525 (2013)

  16. 16.

    et al. Single particle reconstructions of the transferrin-transferrin receptor complex obtained with different specimen preparation techniques. J. Mol. Biol. 355, 1048–1065 (2006)

  17. 17.

    et al. Structural insights into the mechanisms of Mg2+ uptake, transport, and gating by CorA. Proc. Natl Acad. Sci. USA 109, 18459–18464 (2012)

  18. 18.

    et al. Structural asymmetry in the magnesium channel CorA points to sequential allosteric regulation. Proc. Natl Acad. Sci. USA 109, 18809–18814 (2012)

  19. 19.

    et al. Crystal structure of a divalent metal ion transporter CorA at 2.9 angstrom resolution. Science 313, 354–357 (2006)

  20. 20.

    et al. Crystal structure of the CorA Mg2+ transporter. Nature 440, 833–837 (2006)

  21. 21.

    , , & Crystal structure of the calcium release-activated calcium channel Orai. Science 338, 1308–1313 (2012)

  22. 22.

    , & Evolutionary diversity of the mitochondrial calcium uniporter. Science 336, 886 (2012)

  23. 23.

    et al. X-ray structure of the mouse serotonin 5-HT3 receptor. Nature 512, 276–281 (2014)

  24. 24.

    Refined structure of the nicotinic acetylcholine receptor at 4 Å resolution. J. Mol. Biol. 346, 967–989 (2005)

  25. 25.

    , & Predicting coiled coils from protein sequences. Science 252, 1162–1164 (1991)

  26. 26.

    , & A hidden Markov model for predicting transmembrane helices in protein sequences. Proc. Int. Conf. Intell. Syst. Mol. Biol. 6, 175–182 (1998)

  27. 27.

    , , , & HOLE: a program for the analysis of the pore dimensions of ion channel structural models. J. Mol. Graph. 14, 354–360 (1996)

  28. 28.

    , , & Static light scattering to characterize membrane proteins in detergent solution. Methods 46, 73–82 (2008)

  29. 29.

    et al. EMAN2: an extensible image processing suite for electron microscopy. J. Struct. Biol. 157, 38–46 (2007)

  30. 30.

    , & EMAN: semiautomated software for high-resolution single-particle reconstructions. J. Struct. Biol. 128, 82–97 (1999)

  31. 31.

    et al. Structural insight into autoinhibition and histone H3-induced activation of DNMT3A. Nature 517, 640–644 (2015)

  32. 32.

    , & 2D fast rotational matching for image processing of biophysical data. J. Struct. Biol. 144, 51–60 (2003)

  33. 33.

    et al. Fast rotational matching of single-particle images. J. Struct. Biol. 152, 104–112 (2005)

  34. 34.

    et al. NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J. Biomol. NMR 6, 277–293 (1995)

  35. 35.

    et al. The CCPN data model for NMR spectroscopy: development of a software pipeline. Proteins 59, 687–696 (2005)

  36. 36.

    , , , & The program XEASY for computer-supported NMR spectral analysis of biological macromolecules. J. Biomol. NMR 6, 1–10 (1995)

  37. 37.

    , , & Three-dimensional triple resonance NMR spectroscopy of isotopically enriched proteins. J. Magn. Reson. 213, 423–441 (1990)

  38. 38.

    , , & Improved sensitivity and coherence selection for [15N,1H]-TROSY elements in triple resonance experiments. J. Biomol. NMR 15, 181–184 (1999)

  39. 39.

    , , , & Support of 1H NMR assignments in proteins by biosynthetically directed fractional 13C-labeling. J. Biomol. NMR 2, 323–334 (1992)

  40. 40.

    , , & The Xplor-NIH NMR molecular structure determination package. J. Magn. Reson. 160, 65–71 (2003)

  41. 41.

    , , & TALOS+: a hybrid method for predicting protein backbone torsion angles from NMR chemical shifts. J. Biomol. NMR 44, 213–223 (2009)

  42. 42.

    , , & PROCHECK: a program to check the stereochemical quality of protein structures. J. Appl. Crystallogr. 26, 283–291 (1993)

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Acknowledgements

We thank Y. Balazs for helping with ITC measurement and data analysis and Y. Zhang and M. Cao from the EM facility of NCPSS for their assistance with EM data collection. The NMR data were collected at the NMR facility of NCPSS and MIT-Harvard CMR (supported by NIH grant P41 EB-002026). This work was supported by CAS grant XDB08030301 and NIH grant GM094608 to J.J.C. V.K.M. is an Investigator of the Howard Hughes Medical Institute. Y.C. is supported by CAS grant XDB08030201. C.C. is supported by the China Scholarship Council.

Author information

Author notes

    • Kirill Oxenoid
    • , Ying Dong
    • , Chan Cao
    •  & Tanxing Cui

    These authors contributed equally to this work.

Affiliations

  1. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Kirill Oxenoid
    • , Chan Cao
    • , Tanxing Cui
    •  & James J. Chou
  2. State Key Laboratory of Molecular Biology, National Center for Protein Science Shanghai, Shanghai Institute of Biochemistry and Cell Biology, Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 200031, China

    • Ying Dong
    • , Liangliang Kong
    • , Zhijun Liu
    • , Bo Ouyang
    • , Yao Cong
    •  & James J. Chou
  3. State Key Laboratory of Elemento-Organic Chemistry and College of Chemistry, Nankai University, Tianjin 300071, China

    • Chan Cao
  4. Department of Molecular Biology and Howard Hughes Medical Institute, Massachusetts General Hospital, Boston, Massachusetts 02114, USA

    • Yasemin Sancak
    • , Andrew L. Markhard
    • , Zenon Grabarek
    •  & Vamsi K. Mootha

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Contributions

T.C., Y.D., Y.S., C.C., V.K.M. and J.J.C. conceived the study; T.C., Y.S. and C.C. designed protein constructs for structural studies; C.C. and K.O. performed inhibitor binding studies; Y.S., A.L.M. and Z.G. performed structure guided functional experiments and analysis; Y.D., L.K. and Y.C. prepared EM samples and performed EM analysis. K.O., T.C., C.C. and J.J.C. collected NMR data and solved the structure; V.K.M., J.J.C. and K.O. wrote the paper and all authors contributed to editing of the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to James J. Chou.

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https://doi.org/10.1038/nature17656

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