Review Article | Published:

Mitochondrial proteins: from biogenesis to functional networks

Nature Reviews Molecular Cell Biologyvolume 20pages267284 (2019) | Download Citation


Mitochondria are essential for the viability of eukaryotic cells as they perform crucial functions in bioenergetics, metabolism and signalling and have been associated with numerous diseases. Recent functional and proteomic studies have revealed the remarkable complexity of mitochondrial protein organization. Protein machineries with diverse functions such as protein translocation, respiration, metabolite transport, protein quality control and the control of membrane architecture interact with each other in dynamic networks. In this Review, we discuss the emerging role of the mitochondrial protein import machinery as a key organizer of these mitochondrial protein networks. The preprotein translocases that reside on the mitochondrial membranes not only function during organelle biogenesis to deliver newly synthesized proteins to their final mitochondrial destination but also cooperate with numerous other mitochondrial protein complexes that perform a wide range of functions. Moreover, these protein networks form membrane contact sites, for example, with the endoplasmic reticulum, that are key for integration of mitochondria with cellular function, and defects in protein import can lead to diseases.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Additional information

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


  1. 1.

    Neupert, W. A perspective on transport of proteins into mitochondria: a myriad of open questions. J. Mol. Biol. 427, 1135–1158 (2015).

  2. 2.

    Wiedemann, N. & Pfanner, N. Mitochondrial machineries for protein import and assembly. Annu. Rev. Biochem. 86, 685–714 (2017).

  3. 3.

    van der Bliek, A. M., Sedensky, M. M. & Morgan, P. G. Cell biology of the mitochondrion. Genetics 207, 843–871 (2017).

  4. 4.

    Lill, R. Function and biogenesis of iron–sulphur proteins. Nature 460, 831–838 (2009).

  5. 5.

    Ott, M., Amunts, A. & Brown, A. Organization and regulation of mitochondrial protein synthesis. Annu. Rev. Biochem. 85, 77–101 (2016).

  6. 6.

    Hell, K., Neupert, W. & Stuart, R. A. Oxa1p acts as a general membrane insertion machinery for proteins encoded by mitochondrial DNA. EMBO J. 20, 1281–1288 (2001).

  7. 7.

    Labbé, K., Murley, A. & Nunnari, J. Determinants and functions of mitochondrial behavior. Annu. Rev. Cell Dev. Biol. 30, 357–391 (2014).

  8. 8.

    Westermann, B. Mitochondrial fusion and fission in cell life and death. Nat. Rev. Mol. Cell. Biol. 11, 872–884 (2010).

  9. 9.

    von der Malsburg, K. et al. Dual role of mitofilin in mitochondrial membrane organization and protein biogenesis. Dev. Cell 21, 694–707 (2011).

  10. 10.

    Harner, M. et al. The mitochondrial contact site complex, a determinant of mitochondrial architecture. EMBO J. 30, 4356–4370 (2011).

  11. 11.

    Hoppins, S. et al. A mitochondrial-focused genetic interaction map reveals a scaffold-like complex required for inner membrane organization in mitochondria. J. Cell Biol. 195, 323–340 (2011). References 9–11 report the identification of the MICOS, a multisubunit complex that links outer and inner membranes and is crucial for the maintenance of crista junctions.

  12. 12.

    Aaltonen, M. J. et al. MICOS and phospholipid transfer by Ups2–Mdm35 organize membrane lipid synthesis in mitochondria. J. Cell Biol. 213, 525–534 (2016).

  13. 13.

    Ott, C. et al. Sam50 functions in mitochondrial intermembrane space bridging and biogenesis of respiratory complexes. Mol. Cell. Biol. 32, 1173–1188 (2012).

  14. 14.

    Shpilka, T. & Haynes, C. M. The mitochondrial UPR: mechanisms, physiological functions and implications in ageing. Nat. Rev. Mol. Cell. Biol. 19, 109–120 (2018).

  15. 15.

    Pickles, S., Vigié, P. & Youle, R. J. Mitophagy and quality control mechanisms in mitochondrial maintenance. Curr. Biol. 28, R170–R185 (2018).

  16. 16.

    Harper, J. W., Ordureau, A. & Heo, J.-M. Building and decoding ubiquitin chains for mitophagy. Nat. Rev. Mol. Cell. Biol. 19, 93–108 (2018).

  17. 17.

    Cosentino, K. & Garcia-Saez, A. J. Bax and Bak pores: are we closing the circle? Trends Cell Biol. 27, 266–275 (2017).

  18. 18.

    Rugarli, E. I. & Langer, T. Mitochondrial quality control: a matter of life and death for neurons. EMBO J. 31, 1336–1349 (2012).

  19. 19.

    Sena, L. A. & Chandel, N. S. Physiological roles of mitochondrial reactive oxygen species. Mol. Cell 48, 158–167 (2012).

  20. 20.

    Calvo, S. E., Clauser, K. R. & Mootha, V. K. MitoCarta2.0: an updated inventory of mammalian mitochondrial proteins. Nucleic Acids Res. 44, D1251–D1257 (2016).

  21. 21.

    Forner, F., Foster, L. J., Campanaro, S., Valle, G. & Mann, M. Quantitative proteomic comparison of rat mitochondria from muscle, heart, and liver. Mol. Cell. Proteom. 5, 608–619 (2006).

  22. 22.

    Gaucher, S. P. et al. Expanded coverage of the human heart mitochondrial proteome using multidimensional liquid chromatography coupled with tandem mass spectrometry. J. Proteome Res. 3, 495–505 (2004).

  23. 23.

    Hung, V. et al. Proteomic mapping of the human mitochondrial intermembrane space in live cells via ratiometric APEX tagging. Mol. Cell 55, 332–341 (2014).

  24. 24.

    Lefort, N. et al. Proteome profile of functional mitochondria from human skeletal muscle using one-dimensional gel electrophoresis and HPLC-ESI-MS/MS. J. Proteom. 72, 1046–1060 (2009).

  25. 25.

    McDonald, T. et al. Expanding the subproteome of the inner mitochondria using protein separation technologies: one- and two-dimensional liquid chromatography and two-dimensional gel electrophoresis. Mol. Cell. Proteom. 5, 2392–2411 (2006).

  26. 26.

    Morgenstern, M. et al. Definition of a high-confidence mitochondrial proteome at quantitative scale. Cell Rep. 19, 2836–2852 (2017). This is a systematic quantitative analysis of the proteome of yeast mitochondria, revealing the absolute copy numbers of most mitochondrial protein machineries under fermentable and respiratory growth conditions.

  27. 27.

    Ohlmeier, S., Kastaniotis, A. J., Hiltunen, J. K. & Bergmann, U. The yeast mitochondrial proteome, a study of fermentative and respiratory growth. J. Biol. Chem. 279, 3956–3979 (2004).

  28. 28.

    Pagliarini, D. J. et al. A mitochondrial protein compendium elucidates complex I disease biology. Cell 134, 112–123 (2008).

  29. 29.

    Prokisch, H. et al. Integrative analysis of the mitochondrial proteome in yeast. PLOS Biol. 2, e160 (2004).

  30. 30.

    Reinders, J., Zahedi, R. P., Pfanner, N., Meisinger, C. & Sickmann, A. Toward the complete yeast mitochondrial proteome: multidimensional separation techniques for mitochondrial proteomics. J. Proteome Res. 5, 1543–1554 (2006).

  31. 31.

    Rhee, H.-W. et al. Proteomic mapping of mitochondria in living cells via spatially restricted enzymatic tagging. Science 339, 1328–1331 (2013).

  32. 32.

    Sickmann, A. et al. The proteome of Saccharomyces cerevisiae mitochondria. Proc. Natl Acad. Sci. USA 100, 13207–13212 (2003).

  33. 33.

    Smith, A. C. & Robinson, A. J. MitoMinerv3.1, an update on the mitochondrial proteomics database. Nucleic Acids Res. 44, D1258–D1261 (2016).

  34. 34.

    Taylor, S. W. et al. Characterization of the human heart mitochondrial proteome. Nat. Biotechnol. 21, 281–286 (2003).

  35. 35.

    Vögtle, F. N. et al. Landscape of submitochondrial protein distribution. Nat. Commun. 8, 290 (2017).

  36. 36.

    Vögtle, F. N. et al. Intermembrane space proteome of yeast mitochondria. Mol. Cell. Proteom. 11, 1840–1852 (2012).

  37. 37.

    Zahedi, R. P. et al. Proteomic analysis of the yeast mitochondrial outer membrane reveals accumulation of a subclass of preproteins. Mol. Biol. Cell 17, 1436–1450 (2006).

  38. 38.

    Zhang, J. et al. Systematic characterization of the murine mitochondrial proteome using functionally validated cardiac mitochondria. Proteomics 8, 1564–1575 (2008).

  39. 39.

    de Godoy, L. M. F. et al. Comprehensive mass-spectrometry-based proteome quantification of haploid versus diploid yeast. Nature 455, 1251–1254 (2008).

  40. 40.

    Schwanhäusser, B. et al. Global quantification of mammalian gene expression control. Nature 473, 337–342 (2011).

  41. 41.

    Beck, M. et al. The quantitative proteome of a human cell line. Mol. Systems Biol. 7, 549 (2011).

  42. 42.

    Kulak, N. A., Pichler, G., Paron, I., Nagaraj, N. & Mann, M. Minimal, encapsulated proteomic-sample processing applied to copy-number estimation in eukaryotic cells. Nat. Methods 11, 319–324 (2014).

  43. 43.

    Wiśniewski, J. R., Hein, M. Y., Cox, J. & Mann, M. A ‘proteomic ruler’ for protein copy number and concentration estimation without spike-in standards. Mol. Cell. Proteom. 13, 3497–3506 (2014).

  44. 44.

    Paulo, J. A. et al. Quantitative mass spectrometry-based multiplexing compares the abundance of 5000 S. cerevisiae proteins across 10 carbon sources. J. Proteom. 148, 85–93 (2016).

  45. 45.

    Harbauer, A. B. et al. Mitochondria: cell cycle-dependent regulation of mitochondrial preprotein translocase. Science 346, 1109–1113 (2014).

  46. 46.

    Gerbeth, C. et al. Glucose-induced regulation of protein import receptor Tom22 by cytosolic and mitochondria-bound kinases. Cell Metab. 18, 578–587 (2013).

  47. 47.

    Rao, S. et al. Biogenesis of the preprotein translocase of the outer mitochondrial membrane: protein kinase A phosphorylates the precursor of Tom40 and impairs its import. Mol. Biol. Cell 23, 1618–1627 (2012).

  48. 48.

    Schmidt, O. et al. Regulation of mitochondrial protein import by cytosolic kinases. Cell 144, 227–239 (2011). This study reports that biogenesis and activity of the TOM complex are regulated by cytosolic kinases, revealing the main protein import site of mitochondria as a major target for cytosolic signalling pathways.

  49. 49.

    Vögtle, F. N. et al. Global analysis of the mitochondrial N-proteome identifies a processing peptidase critical for protein stability. Cell 139, 428–439 (2009).

  50. 50.

    Abe, Y. et al. Structural basis of presequence recognition by the mitochondrial protein import receptor Tom20. Cell 100, 551–560 (2000).

  51. 51.

    van Wilpe, S. et al. Tom22 is a multifunctional organizer of the mitochondrial preprotein translocase. Nature 401, 485–489 (1999).

  52. 52.

    Kuszak, A. J. et al. Evidence of distinct channel conformations and substrate binding affinities for the mitochondrial outer membrane protein translocase pore Tom40. J. Biol. Chem. 290, 26204–26217 (2015).

  53. 53.

    Melin, J. et al. Presequence recognition by the Tom40 channel contributes to precursor translocation into the mitochondrial matrix. Mol. Cell. Biol. 34, 3473–3485 (2014).

  54. 54.

    Lohret, T. A., Jensen, R. E. & Kinnally, K. W. Tim23, a protein import component of the mitochondrial inner membrane, is required for normal activity of the multiple conductance channel, MCC. J. Cell Biol. 137, 377–386 (1997).

  55. 55.

    Bauer, M. F., Sirrenberg, C., Neupert, W. & Brunner, M. Role of Tim23 as voltage sensor and presequence receptor in protein import into mitochondria. Cell 87, 33–41 (1996).

  56. 56.

    Dekker, P. J. et al. Identification of MIM23, a putative component of the protein import machinery of the mitochondrial inner membrane. FEBS Lett. 330, 66–70 (1993).

  57. 57.

    Demishtein-Zohary, K., Marom, M., Neupert, W., Mokranjac, D. & Azem, A. GxxxG motifs hold the TIM23 complex together. FEBS J. 282, 2178–2186 (2015).

  58. 58.

    Truscott, K. N. et al. A presequence- and voltage-sensitive channel of the mitochondrial preprotein translocase formed by Tim23. Nat. Struct. Biol. 8, 1074–1082 (2001).

  59. 59.

    Malhotra, K. et al. Cardiolipin mediates membrane and channel interactions of the mitochondrial TIM23 protein import complex receptor Tim50. Sci. Adv. 3, e1700532 (2017).

  60. 60.

    Denkert, N. et al. Cation selectivity of the presequence translocase channel Tim23 is crucial for efficient protein import. eLife 6, e28324 (2017).

  61. 61.

    Ramesh, A. et al. A disulfide bond in the TIM23 complex is crucial for voltage gating and mitochondrial protein import. J. Cell Biol. 214, 417–431 (2016).

  62. 62.

    Kang, P. J. et al. Requirement for hsp70 in the mitochondrial matrix for translocation and folding of precursor proteins. Nature 348, 137–143 (1990).

  63. 63.

    Demishtein-Zohary, K. et al. Role of Tim17 in coupling the import motor to the translocation channel of the mitochondrial presequence translocase. eLife 6, e22696 (2017).

  64. 64.

    Ting, S.-Y., Yan, N. L., Schilke, B. A. & Craig, E. A. Dual interaction of scaffold protein Tim44 of mitochondrial import motor with channel-forming translocase subunit Tim23. eL ife 6, e23609 (2017).

  65. 65.

    Banerjee, R., Gladkova, C., Mapa, K., Witte, G. & Mokranjac, D. Protein translocation channel of mitochondrial inner membrane and matrix-exposed import motor communicate via two-domain coupling protein. eLife 4, e11897 (2015).

  66. 66.

    Sikor, M., Mapa, K., von Voithenberg, L. V., Mokranjac, D. & Lamb, D. C. Real-time observation of the conformational dynamics of mitochondrial Hsp70 by spFRET. EMBO J. 32, 1639–1649 (2013).

  67. 67.

    Schendzielorz, A. B. et al. Two distinct membrane potential-dependent steps drive mitochondrial matrix protein translocation. J. Cell Biol. 216, 83–92 (2017).

  68. 68.

    Schulz, C. & Rehling, P. Remodelling of the active presequence translocase drives motor-dependent mitochondrial protein translocation. Nat. Commun. 5, 4349 (2014).

  69. 69.

    Fukasawa, Y. et al. MitoFates: improved prediction of mitochondrial targeting sequences and their cleavage sites. Mol. Cell. Proteom. 14, 1113–1126 (2015).

  70. 70.

    Varshavsky, A. The N-end rule pathway and regulation by proteolysis. Protein Sci. 20, 1298–1345 (2011).

  71. 71.

    Veling, M. T. et al. Multi-omic mitoprotease profiling defines a role for Oct1p in coenzyme Q production. Mol. Cell 68, 970–977 (2017).

  72. 72.

    Vögtle, F. N. et al. Mitochondrial protein turnover: role of the precursor intermediate peptidase Oct1 in protein stabilization. Mol. Biol. Cell 22, 2135–2143 (2011).

  73. 73.

    Cheng, M. Y. et al. Mitochondrial heat-shock protein hsp60 is essential for assembly of proteins imported into yeast mitochondria. Nature 337, 620–625 (1989).

  74. 74.

    Ostermann, J., Horwich, A. L., Neupert, W. & Hartl, F. U. Protein folding in mitochondria requires complex formation with hsp60 and ATP hydrolysis. Nature 341, 125–130 (1989).

  75. 75.

    Ieva, R. et al. Mgr2 functions as lateral gatekeeper for preprotein sorting in the mitochondrial inner membrane. Mol. Cell 56, 641–652 (2014). This paper identifies a lateral gatekeeper protein at the TIM23 complex that controls the proper release of preproteins with hydrophobic sorting signals into the inner membrane.

  76. 76.

    Schendzielorz, A. B. et al. Motor recruitment to the TIM23 channel’s lateral gate restricts polypeptide release into the inner membrane. Nat. Commun. 9, 4028 (2018).

  77. 77.

    Stiller, S. B. et al. Mitochondrial OXA translocase plays a major role in biogenesis of inner-membrane proteins. Cell Metab. 23, 901–908 (2016).

  78. 78.

    Park, K., Botelho, S. C., Hong, J., Osterberg, M. & Kim, H. Dissecting stop transfer versus conservative sorting pathways for mitochondrial inner membrane proteins in vivo. J. Biol. Chem. 288, 1521–1532 (2013).

  79. 79.

    Bohnert, M. et al. Cooperation of stop-transfer and conservative sorting mechanisms in mitochondrial protein transport. Curr. Biol. 20, 1227–1232 (2010).

  80. 80.

    Herrmann, J. M., Neupert, W. & Stuart, R. A. Insertion into the mitochondrial inner membrane of a polytopic protein, the nuclear-encoded Oxa1p. EMBO J. 16, 2217–2226 (1997).

  81. 81.

    Bausewein, T. et al. Cryo-EM structure of the TOM core complex from Neurospora crassa. Cell 170, 693–700 (2017). This paper provides a cryoelectron microscopy structure of the TOM core complex of the mitochondrial outer membrane. Two translocation pores formed by Tom40 β-barrels are connected by two copies of the central receptor Tom22.

  82. 82.

    Shiota, T. et al. Molecular architecture of the active mitochondrial protein gate. Science 349, 1544–1548 (2015). This study maps the TOM complex architecture by site-specific crosslinking in the native membrane, revealing different translocation pathways for hydrophilic and hydrophobic precursor proteins through the Tom40 channel.

  83. 83.

    Backes, S. et al. Tom70 enhances mitochondrial preprotein import efficiency by binding to internal targeting sequences. J. Cell Biol. 217, 1369–1382 (2018). This study reports a role of Tom70 in the import of presequence-containing preproteins that contain additional internal targeting signals. While Tom20 and Tom22 bind the amino-terminal presequences, Tom70 binds the internal signals to prevent aggregation of the preproteins.

  84. 84.

    Melin, J. et al. A presequence-binding groove in Tom70 supports import of Mdl1 into mitochondria. Biochim. Biophys. Acta 1853, 1850–1859 (2015).

  85. 85.

    Yamamoto, H. et al. Roles of Tom70 in Import of presequence-containing mitochondrial proteins. J. Biol. Chem. 284, 31635–31646 (2009).

  86. 86.

    Young, J. C., Hoogenraad, N. J. & Hartl, F. U. Molecular chaperones Hsp90 and Hsp70 deliver preproteins to the mitochondrial import receptor Tom70. Cell 112, 41–50 (2003).

  87. 87.

    Wiedemann, N., Pfanner, N. & Ryan, M. T. The three modules of ADP/ATP carrier cooperate in receptor recruitment and translocation into mitochondria. EMBO J. 20, 951–960 (2001).

  88. 88.

    Curran, S. P., Leuenberger, D., Schmidt, E. & Koehler, C. M. The role of the Tim8p-Tim13p complex in a conserved import pathway for mitochondrial polytopic inner membrane proteins. J. Cell Biol. 158, 1017–1027 (2002).

  89. 89.

    Vial, S. et al. Assembly of Tim9 and Tim10 into a functional chaperone. J. Biol. Chem. 277, 36100–36108 (2002).

  90. 90.

    Koehler, C. M. et al. Tim9p, an essential partner subunit of Tim10p for the import of mitochondrial carrier proteins. EMBO J. 17, 6477–6486 (1998).

  91. 91.

    Sirrenberg, C. et al. Carrier protein import into mitochondria mediated by the intermembrane proteins Tim10/Mrs11 and Tim12/Mrs5. Nature 391, 912–915 (1998).

  92. 92.

    Weinhäupl, K. et al. Structural basis of membrane protein chaperoning through the mitochondrial intermembrane space. Cell 175, 1365–1379 (2018).

  93. 93.

    Okamoto, H., Miyagawa, A., Shiota, T., Tamura, Y. & Endo, T. Intramolecular disulfide bond of Tim22 protein maintains integrity of the TIM22 complex in the mitochondrial inner membrane. J. Biol. Chem. 289, 4827–4838 (2014).

  94. 94.

    Wrobel, L., Trojanowska, A., Sztolsztener, M. E. & Chacinska, A. Mitochondrial protein import: Mia40 facilitates Tim22 translocation into the inner membrane of mitochondria. Mol. Biol. Cell 24, 543–554 (2013).

  95. 95.

    Rehling, P. et al. Protein insertion into the mitochondrial inner membrane by a twin-pore translocase. Science 299, 1747–1751 (2003).

  96. 96.

    Kerscher, O., Holder, J., Srinivasan, M., Leung, R. S. & Jensen, R. E. The Tim54p-Tim22p complex mediates insertion of proteins into the mitochondrial inner membrane. J. Cell Biol. 139, 1663–1675 (1997).

  97. 97.

    Sirrenberg, C., Bauer, M. F., Guiard, B., Neupert, W. & Brunner, M. Import of carrier proteins into the mitochondrial inner membrane mediated by Tim22. Nature 384, 582–585 (1996).

  98. 98.

    Koch, J. R. & Schmid, F. X. Mia40 combines thiol oxidase and disulfide isomerase activity to efficiently catalyze oxidative folding in mitochondria. J. Mol. Biol. 426, 4087–4098 (2014).

  99. 99.

    Chacinska, A. et al. Essential role of Mia40 in import and assembly of mitochondrial intermembrane space proteins. EMBO J. 23, 3735–3746 (2004).

  100. 100.

    Mesecke, N. et al. A disulfide relay system in the intermembrane space of mitochondria that mediates protein import. Cell 121, 1059–1069 (2005).

  101. 101.

    Milenkovic, D. et al. Identification of the signal directing Tim9 and Tim10 into the intermembrane space of mitochondria. Mol. Biol. Cell 20, 2530–2539 (2009).

  102. 102.

    Sideris, D. P. et al. A novel intermembrane space-targeting signal docks cysteines onto Mia40 during mitochondrial oxidative folding. J. Cell Biol. 187, 1007–1022 (2009).

  103. 103.

    Peleh, V., Cordat, E. & Herrmann, J. M. Mia40 is a trans-site receptor that drives protein import into the mitochondrial intermembrane space by hydrophobic substrate binding. eLife 5, e16177 (2016).

  104. 104.

    Neal, S. E. et al. Mia40 protein serves as an electron sink in the Mia40-Erv1 import pathway. J. Biol. Chem. 290, 20804–20814 (2015).

  105. 105.

    Kojer, K., Peleh, V., Calabrese, G., Herrmann, J. M. & Riemer, J. Kinetic control by limiting glutaredoxin amounts enables thiol oxidation in the reducing mitochondrial intermembrane space. Mol. Biol. Cell 26, 195–204 (2015).

  106. 106.

    Jores, T. et al. Characterization of the targeting signal in mitochondrial β-barrel proteins. Nat. Commun. 7, 12036 (2016).

  107. 107.

    Klein, A. et al. Characterization of the insertase for β-barrel proteins of the outer mitochondrial membrane. J. Cell Biol. 199, 599–611 (2012).

  108. 108.

    Wiedemann, N. et al. Machinery for protein sorting and assembly in the mitochondrial outer membrane. Nature 424, 565–571 (2003).

  109. 109.

    Paschen, S. A. et al. Evolutionary conservation of biogenesis of beta-barrel membrane proteins. Nature 426, 862–866 (2003).

  110. 110.

    Höhr, A. I. C. et al. Membrane protein insertion through a mitochondrial β-barrel gate. Science 359, eaah6834 (2018). This study maps the membrane insertion pathway of β-barrel precursors through the SAM channel, signal-induced opening of the lateral gate of Sam50 and release of the folded β-barrel protein into the outer membrane.

  111. 111.

    Kutik, S. et al. Dissecting membrane insertion of mitochondrial β-barrel proteins. Cell 132, 1011–1024 (2008).

  112. 112.

    Krüger, V. et al. Identification of new channels by systematic analysis of the mitochondrial outer membrane. J. Cell Biol. 216, 3485–3495 (2017).

  113. 113.

    Dimmer, K. S. et al. A crucial role for Mim2 in the biogenesis of mitochondrial outer membrane proteins. J. Cell Sci. 125, 3464–3473 (2012).

  114. 114.

    Papic, D., Krumpe, K., Dukanovic, J., Dimmer, K. S. & Rapaport, D. Multispan mitochondrial outer membrane protein Ugo1 follows a unique Mim1-dependent import pathway. J. Cell Biol. 194, 397–405 (2011).

  115. 115.

    Hulett, J. M. et al. The transmembrane segment of Tom20 is recognized by Mim1 for docking to the mitochondrial TOM complex. J. Mol. Biol. 376, 694–704 (2008).

  116. 116.

    Popov-Čeleketić, J., Waizenegger, T. & Rapaport, D. Mim1 functions in an oligomeric form to facilitate the integration of Tom20 into the mitochondrial outer membrane. J. Mol. Biol. 376, 671–680 (2008).

  117. 117.

    Becker, T. et al. The mitochondrial import protein Mim1 promotes biogenesis of multispanning outer membrane proteins. J. Cell Biol. 194, 387–395 (2011).

  118. 118.

    Dukanovic, J. & Rapaport, D. Multiple pathways in the integration of proteins into the mitochondrial outer membrane. Biochim. Biophys. Acta 1808, 971–980 (2011).

  119. 119.

    Keskin, A., Akdoğan, E. & Dunn, C. D. Evidence for amino acid snorkeling from a high-resolution, in vivo analysis of Fis1 tail-anchor insertion at the mitochondrial outer membrane. Genetics 205, 691–705 (2017).

  120. 120.

    Vögtle, F. N. et al. The fusogenic lipid phosphatidic acid promotes the biogenesis of mitochondrial outer membrane protein Ugo1. J. Cell Biol. 210, 951–960 (2015).

  121. 121.

    Sauerwald, J. et al. Genome-wide screens in Saccharomyces cerevisiae highlight a role for cardiolipin in biogenesis of mitochondrial outer membrane multispan proteins. Mol. Cell. Biol. 35, 3200–3211 (2015).

  122. 122.

    Albrecht, R. et al. The Tim21 binding domain connects the preprotein translocases of both mitochondrial membranes. EMBO Rep. 7, 1233–1238 (2006).

  123. 123.

    Chacinska, A. et al. Distinct forms of mitochondrial TOM-TIM supercomplexes define signal-dependent states of preprotein sorting. Mol. Cell. Biol. 30, 307–318 (2010).

  124. 124.

    Gold, V. A. M. et al. Visualizing active membrane protein complexes by electron cryotomography. Nat. Commun. 5, 4129 (2014).

  125. 125.

    Qiu, J. et al. Coupling of mitochondrial import and export translocases by receptor-mediated supercomplex formation. Cell 154, 596–608 (2013). This study identifies a TOM–SAM supercomplex that facilitates the efficient transfer of β-barrel precursors from the TOM import channel to the SAM membrane insertion sites of the mitochondrial outer membrane.

  126. 126.

    Waegemann, K., Popov-Čeleketić, D., Neupert, W., Azem, A. & Mokranjac, D. Cooperation of TOM and TIM23 complexes during translocation of proteins into mitochondria. J. Mol. Biol. 427, 1075–1084 (2015).

  127. 127.

    Wenz, L.-S. et al. Sam37 is crucial for formation of the mitochondrial TOM-SAM supercomplex, thereby promoting β-barrel biogenesis. J. Cell Biol. 210, 1047–1054 (2015).

  128. 128.

    Callegari, S. et al. TIM29 is a subunit of the human carrier translocase required for protein transport. FEBS Lett. 590, 4147–4158 (2016).

  129. 129.

    Kang, Y. et al. Tim29 is a novel subunit of the human TIM22 translocase and is involved in complex assembly and stability. eL ife 5, e17463 (2016).

  130. 130.

    Gornicka, A. et al. A discrete pathway for the transfer of intermembrane space proteins across the outer membrane of mitochondria. Mol. Biol. Cell 25, 3999–4009 (2014).

  131. 131.

    Wiedemann, N. et al. Biogenesis of yeast mitochondrial cytochrome c: a unique relationship to the TOM machinery. J. Mol. Biol. 327, 465–474 (2003).

  132. 132.

    Sinha, D., Srivastava, S., Krishna, L. & D’Silva, P. Unraveling the intricate organization of mammalian mitochondrial presequence translocases: existence of multiple translocases for maintenance of mitochondrial function. Mol. Cell. Biol. 34, 1757–1775 (2014).

  133. 133.

    Rainbolt, T. K., Atanassova, N., Genereux, J. C. & Wiseman, R. L. Stress-regulated translational attenuation adapts mitochondrial protein import through Tim17A degradation. Cell Metab. 18, 908–919 (2013).

  134. 134.

    Opalińska, M., Parys, K., Murcha, M. W. & Jańska, H. The plant i-AAA protease controls the turnover of an essential mitochondrial protein import component. J. Cell Sci. 131, jcs200733 (2018).

  135. 135.

    Wenz, L.-S. et al. The presequence pathway is involved in protein sorting to the mitochondrial outer membrane. EMBO Rep. 15, 678–685 (2014).

  136. 136.

    Song, J., Tamura, Y., Yoshihisa, T. & Endo, T. A novel import route for an N-anchor mitochondrial outer membrane protein aided by the TIM23 complex. EMBO Rep. 15, 670–677 (2014).

  137. 137.

    Lee, C. M., Sedman, J., Neupert, W. & Stuart, R. A. The DNA helicase, Hmi1p, is transported into mitochondria by a C-terminal cleavable targeting signal. J. Biol. Chem. 274, 20937–20942 (1999).

  138. 138.

    Ieva, R. et al. Mitochondrial inner membrane protease promotes assembly of presequence translocase by removing a carboxy-terminal targeting sequence. Nat. Commun. 4, 2853 (2013).

  139. 139.

    Sinzel, M. et al. Mcp3 is a novel mitochondrial outer membrane protein that follows a unique IMP-dependent biogenesis pathway. EMBO Rep. 17, 965–981 (2016).

  140. 140.

    Acin-Perez, R., Fernández-Silva, P., Peleato, M. L., Pérez-Martos, A. & Enríquez, J. A. Respiratory active mitochondrial supercomplexes. Mol. Cell 32, 529–539 (2008).

  141. 141.

    Melber, A. & Winge, D. R. Inner secrets of the respirasome. Cell 167, 1450–1452 (2016).

  142. 142.

    Wu, M., Gu, J., Guo, R., Huang, Y. & Yang, M. Structure of mammalian respiratory supercomplex I1III2IV1. Cell 167, 1598–1609 (2016). This study provides a cryoelectron microscopy structure of the 1.7 MDa respiratory supercomplex at near-atomic resolution, revealing the arrangement of complexes I, III and IV and the position of cofactors and phospholipids.

  143. 143.

    Milenkovic, D., Blaza, J. N., Larsson, N.-G. & Hirst, J. The enigma of the respiratory chain supercomplex. Cell Metab. 25, 765–776 (2017).

  144. 144.

    Schweppe, D. K. et al. Mitochondrial protein interactome elucidated by chemical cross-linking mass spectrometry. Proc. Natl Acad. Sci. USA 114, 1732–1737 (2017).

  145. 145.

    Chen, Y.-C. et al. Identification of a protein mediating respiratory supercomplex stability. Cell Metab. 15, 348–360 (2012).

  146. 146.

    Strogolova, V., Furness, A., Robb-McGrath, M., Garlich, J. & Stuart, R. A. Rcf1 and Rcf2, members of the hypoxia-induced gene 1 protein family, are critical components of the mitochondrial cytochrome bc 1-cytochrome c oxidase supercomplex. Mol. Cell. Biol. 32, 1363–1373 (2012).

  147. 147.

    Vukotic, M. et al. Rcf1 mediates cytochrome oxidase assembly and respirasome formation, revealing heterogeneity of the enzyme complex. Cell Metab. 15, 336–347 (2012).

  148. 148.

    Singhal, R. K. et al. Coi1 is a novel assembly factor of the yeast complex III-complex IV supercomplex. Mol. Biol. Cell 28, 2609–2622 (2017).

  149. 149.

    Dannenmaier, S. et al. Complete native stable isotope labeling by amino acids of Saccharomyces cerevisiae for global proteomic analysis. Anal. Chem. 90, 10501–10509 (2018).

  150. 150.

    van der Laan, M. et al. A role for Tim21 in membrane-potential-dependent preprotein sorting in mitochondria. Curr. Biol. 16, 2271–2276 (2006).

  151. 151.

    Wiedemann, N., van der Laan, M., Hutu, D. P., Rehling, P. & Pfanner, N. Sorting switch of mitochondrial presequence translocase involves coupling of motor module to respiratory chain. J. Cell Biol. 179, 1115–1122 (2007).

  152. 152.

    Mehnert, C. S. et al. The mitochondrial ADP/ATP carrier associates with the inner membrane presequence translocase in a stoichiometric manner. J. Biol. Chem. 289, 27352–27362 (2014).

  153. 153.

    Dennerlein, S. et al. MITRAC7 acts as a COX1-specific chaperone and reveals a checkpoint during cytochrome c oxidase assembly. Cell Rep. 12, 1644–1655 (2015).

  154. 154.

    Mick, D. U. et al. MITRAC links mitochondrial protein translocation to respiratory-chain assembly and translational regulation. Cell 151, 1528–1541 (2012).

  155. 155.

    Richter-Dennerlein, R. et al. Mitochondrial protein synthesis adapts to influx of nuclear-encoded protein. Cell 167, 471–483 (2016). This paper identifies a mechanism by which (MITRAC) assembly factors adjust the efficiency of mitochondrial synthesis of membrane-integrated respiratory chain subunits to the import of nuclear-encoded partner proteins, which is termed mitochondrial translational plasticity.

  156. 156.

    Stoldt, S. et al. Spatial orchestration of mitochondrial translation and OXPHOS complex assembly. Nat. Cell Biol. 20, 528–534 (2018).

  157. 157.

    Topf, U. et al. Quantitative proteomics identifies redox switches for global translation modulation by mitochondrially produced reactive oxygen species. Nat. Commun. 9, 324 (2018).

  158. 158.

    Floyd, B. J. et al. Mitochondrial protein interaction mapping identifies regulators of respiratory chain function. Mol. Cell 63, 621–632 (2016).

  159. 159.

    Stefely, J. A. et al. Mitochondrial protein functions elucidated by multi-omic mass spectrometry profiling. Nat. Biotechnol. 34, 1191–1197 (2016).

  160. 160.

    Böttinger, L. et al. Respiratory chain supercomplexes associate with the cysteine desulfurase complex of the iron-sulfur cluster assembly machinery. Mol. Biol. Cell 29, 776–785 (2018).

  161. 161.

    Gerdes, F., Tatsuta, T. & Langer, T. Mitochondrial AAA proteases—towards a molecular understanding of membrane-bound proteolytic machines. Biochim. Biophys. Acta 1823, 49–55 (2012).

  162. 162.

    Puchades, C. et al. Structure of the mitochondrial inner membrane AAA+protease YME1 gives insight into substrate processing. Science 358, eaao0464 (2017).

  163. 163.

    Wu, X., Li, L. & Jiang, H. Mitochondrial inner-membrane protease Yme1 degrades outer-membrane proteins Tom22 and Om45. J. Cell Biol. 217, 139–149 (2018).

  164. 164.

    van der Laan, M., Bohnert, M., Wiedemann, N. & Pfanner, N. Role of MINOS in mitochondrial membrane architecture and biogenesis. Trends Cell Biol. 22, 185–192 (2012).

  165. 165.

    Bohnert, M. et al. Role of mitochondrial inner membrane organizing system in protein biogenesis of the mitochondrial outer membrane. Mol. Biol. Cell 23, 3948–3956 (2012).

  166. 166.

    Zerbes, R. M. et al. Role of MINOS in mitochondrial membrane architecture: cristae morphology and outer membrane interactions differentially depend on mitofilin domains. J. Mol. Biol. 422, 183–191 (2012).

  167. 167.

    Körner, C. et al. The C-terminal domain of Fcj1 is required for formation of crista junctions and interacts with the TOB/SAM complex in mitochondria. Mol. Biol. Cell 23, 2143–2155 (2012).

  168. 168.

    Xie, J., Marusich, M. F., Souda, P., Whitelegge, J. & Capaldi, R. A. The mitochondrial inner membrane protein Mitofilin exists as a complex with SAM50, metaxins 1 and 2, coiled-coil-helix coiled-coil-helix domain-containing protein 3 and 6 and DnaJC11. FEBS Lett. 581, 3545–3549 (2007).

  169. 169.

    Rabl, R. et al. Formation of cristae and crista junctions in mitochondria depends on antagonism between Fcj1 and Su e/g. J. Cell Biol. 185, 1047–1063 (2009).

  170. 170.

    Jans, D. C. et al. STED super-resolution microscopy reveals an array of MINOS clusters along human mitochondria. Proc. Natl Acad. Sci. USA 110, 8936–8941 (2013).

  171. 171.

    Barbot, M. et al. Mic10 oligomerizes to bend mitochondrial inner membranes at cristae junctions. Cell Metab. 21, 756–763 (2015).

  172. 172.

    Bohnert, M. et al. Central role of Mic10 in the mitochondrial contact site and cristae organizing system. Cell Metab. 21, 747–755 (2015).

  173. 173.

    Rampelt, H. et al. Mic10, a core subunit of the mitochondrial contact site and cristae organizing system, interacts with the dimeric F1F0-ATP synthase. J. Mol. Biol. 429, 1162–1170 (2017).

  174. 174.

    Eydt, K., Davies, K. M., Behrendt, C., Wittig, I. & Reichert, A. S. Cristae architecture is determined by an interplay of the MICOS complex and the F1F0 ATP synthase via Mic27 and Mic10. Microb. Cell 4, 259–272 (2017).

  175. 175.

    Friedman, J. R., Mourier, A., Yamada, J., McCaffery, J. M. & Nunnari, J. MICOS coordinates with respiratory complexes and lipids to establish mitochondrial inner membrane architecture. Elife 4, e07739 (2015).

  176. 176.

    Rampelt, H. et al. Assembly of the mitochondrial cristae organizer Mic10 is regulated by Mic26-Mic27 antagonism and cardiolipin. J. Mol. Biol. 430, 1883–1890 (2018).

  177. 177.

    Barrera, M., Koob, S., Dikov, D., Vogel, F. & Reichert, A. S. OPA1 functionally interacts with MIC60 but is dispensable for crista junction formation. FEBS Lett. 590, 3309–3322 (2016).

  178. 178.

    Glytsou, C. et al. Optic atrophy 1 is epistatic to the core MICOS component MIC60 in mitochondrial cristae shape control. Cell Rep. 17, 3024–3034 (2016).

  179. 179.

    Itoh, K., Tamura, Y., Iijima, M. & Sesaki, H. Effects of Fcj1-Mos1 and mitochondrial division on aggregation of mitochondrial DNA nucleoids and organelle morphology. Mol. Biol. Cell 24, 1842–1851 (2013).

  180. 180.

    Li, H. et al. Mic60/Mitofilin determines MICOS assembly essential for mitochondrial dynamics and mtDNA nucleoid organization. Cell Death Differ. 23, 380–392 (2016).

  181. 181.

    Kornmann, B. et al. An ER-mitochondria tethering complex revealed by a synthetic biology screen. Science 325, 477–481 (2009). This paper identifies the ERMES. ERMES-mediated contact sites between ER and mitochondria are involved in lipid transfer and maintenance of mitochondrial morphology.

  182. 182.

    Meisinger, C. et al. The mitochondrial morphology protein Mdm10 functions in assembly of the preprotein translocase of the outer membrane. Dev. Cell 7, 61–71 (2004).

  183. 183.

    Yamano, K., Tanaka-Yamano, S. & Endo, T. Mdm10 as a dynamic constituent of the TOB/SAM complex directs coordinated assembly of Tom40. EMBO Rep. 11, 187–193 (2010).

  184. 184.

    Flinner, N. et al. Mdm10 is an ancient eukaryotic porin co-occurring with the ERMES complex. Biochim. Biophys. Acta 1833, 3314–3325 (2013).

  185. 185.

    Ellenrieder, L. et al. Separating mitochondrial protein assembly and endoplasmic reticulum tethering by selective coupling of Mdm10. Nat. Commun. 7, 13021 (2016).

  186. 186.

    Meisinger, C. et al. Mitochondrial protein sorting: differentiation of beta-barrel assembly by Tom7-mediated segregation of Mdm10. J. Biol. Chem. 281, 22819–22826 (2006).

  187. 187.

    Stroud, D. A. et al. Composition and topology of the endoplasmic reticulum-mitochondria encounter structure. J. Mol. Biol. 413, 743–750 (2011).

  188. 188.

    Yamano, K., Tanaka-Yamano, S. & Endo, T. Tom7 regulates Mdm10-mediated assembly of the mitochondrial import channel protein Tom40. J. Biol. Chem. 285, 41222–41231 (2010).

  189. 189.

    Müller, C. S. et al. Cryo-slicing blue native-mass spectrometry (csBN-MS), a novel technology for high resolution complexome profiling. Mol. Cell. Proteom. 15, 669–681 (2016).

  190. 190.

    Elbaz-Alon, Y. et al. Lam6 regulates the extent of contacts between organelles. Cell Rep. 12, 7–14 (2015).

  191. 191.

    Murley, A. et al. Ltc1 is an ER-localized sterol transporter and a component of ER-mitochondria and ER-vacuole contacts. J. Cell Biol. 209, 539–548 (2015). References 190 and 191 report that the lipid transfer protein Lam6 is located at and regulates contact sites between ER, mitochondria and further organelles. Lam6 interacts with the receptor Tom70 in ER–mitochondria contact sites.

  192. 192.

    Filadi, R. et al. TOM70 sustains cell bioenergetics by promoting IP3R3-mediated ER to mitochondria Ca2+ transfer. Curr. Biol. 28, 369–382 (2018). This paper shows that the receptor TOM70 of the mitochondrial outer membrane interacts with inositol trisphosphate receptors of the ER, supporting the formation of contact sites for Ca 2+ transfer to mitochondria.

  193. 193.

    González Montoro, A. et al. Vps39 interacts with Tom40 to establish one of two functionally distinct vacuole-mitochondria contact sites. Dev. Cell 45, 621–636 (2018).

  194. 194.

    McLelland, G.-L., Lee, S. A., McBride, H. M. & Fon, E. A. Syntaxin-17 delivers PINK1/parkin-dependent mitochondrial vesicles to the endolysosomal system. J. Cell Biol. 214, 275–291 (2016).

  195. 195.

    Soubannier, V., Rippstein, P., Kaufman, B. A., Shoubridge, E. A. & McBride, H. M. Reconstitution of mitochondria derived vesicle formation demonstrates selective enrichment of oxidized cargo. PLOS ONE 7, e52830 (2012).

  196. 196.

    Soubannier, V. et al. A vesicular transport pathway shuttles cargo from mitochondria to lysosomes. Curr. Biol. 22, 135–141 (2012). References 195 and 196 report that stress conditions can induce the formation of mitochondria-derived vesicles that transport selected cargo such as oxidized proteins to lysosomes as part of a mitochondrial quality control system.

  197. 197.

    Hughes, A. L., Hughes, C. E., Henderson, K. A., Yazvenko, N. & Gottschling, D. E. Selective sorting and destruction of mitochondrial membrane proteins in aged yeast. Elife 5, e13943 (2016).

  198. 198.

    Chen, Y.-C. et al. Msp1/ATAD1 maintains mitochondrial function by facilitating the degradation of mislocalized tail-anchored proteins. EMBO J. 33, 1548–1564 (2014).

  199. 199.

    Okreglak, V. & Walter, P. The conserved AAA-ATPase Msp1 confers organelle specificity to tail-anchored proteins. Proc. Natl Acad. Sci. USA 111, 8019–8024 (2014).

  200. 200.

    Weidberg, H. & Amon, A. MitoCPR — a surveillance pathway that protects mitochondria in response to protein import stress. Science 360, eaan4146 (2018). References 198, 199 and 200 report that the AAA-type ATPase Msp1 extracts mistargeted or non-imported proteins from the mitochondrial outer membrane for degradation by the proteasome in the cytosol.

  201. 201.

    Sekine, S. & Youle, R. J. PINK1 import regulation; a fine system to convey mitochondrial stress to the cytosol. BMC Biol. 16, 2 (2018).

  202. 202.

    Okamoto, K. Quality control: organellophagy: eliminating cellular building blocks via selective autophagy. J. Cell Biol. 205, 435–445 (2014).

  203. 203.

    Suomalainen, A. & Battersby, B. J. Mitochondrial diseases: the contribution of organelle stress responses to pathology. Nat. Rev. Mol. Cell. Biol. 19, 77–92 (2018).

  204. 204.

    Frazier, A. E., Thorburn, D. R. & Compton, A. G. Mitochondrial energy generation disorders: genes, mechanisms and clues to pathology. J. Biol. Chem. (2017).

  205. 205.

    Melber, A. & Haynes, C. M. UPRmt regulation and output: a stress response mediated by mitochondrial-nuclear communication. Cell Res. 28, 281–295 (2018).

  206. 206.

    Nargund, A. M., Pellegrino, M. W., Fiorese, C. J., Baker, B. M. & Haynes, C. M. Mitochondrial import efficiency of ATFS-1 regulates mitochondrial UPR activation. Science 337, 587–590 (2012). Impaired mitochondrial protein import activates an unfolded protein response. The transcription factor ATFS-1 is normally imported into mitochondria and degraded. Disturbance of mitochondrial import leads to cytosolic accumulation of ATFS-1 and translocation into the nucleus, where it induces expression of chaperones and further rescue factors.

  207. 207.

    Fiorese, C. J. et al. The transcription factor ATF5 mediates a mammalian mitochondrial UPR. Curr. Biol. 26, 2037–2043 (2016).

  208. 208.

    Wrobel, L. et al. Mistargeted mitochondrial proteins activate a proteostatic response in the cytosol. Nature 524, 485–488 (2015).

  209. 209.

    Wang, X. & Chen, X. J. A cytosolic network suppressing mitochondria-mediated proteostatic stress and cell death. Nature 524, 481–484 (2015). References 208 and 209 identify a stress response that is induced by accumulation of mitochondrial precursor proteins in the cytosol. The response leads to a decrease in cytosolic protein synthesis and increased proteasomal activity to reduce the accumulation of toxic mistargeted proteins.

  210. 210.

    Bragoszewski, P. et al. Retro-translocation of mitochondrial intermembrane space proteins. Proc. Natl Acad. Sci. USA 112, 7713–7718 (2015).

  211. 211.

    Bragoszewski, P., Gornicka, A., Sztolsztener, M. E. & Chacinska, A. The ubiquitin-proteasome system regulates mitochondrial intermembrane space proteins. Mol. Cell. Biol. 33, 2136–2148 (2013).

  212. 212.

    Kowalski, L. et al. Determinants of the cytosolic turnover of mitochondrial intermembrane space proteins. BMC Biol. 16, 66 (2018).

  213. 213.

    Akabane, S. et al. PKA regulates PINK1 stability and Parkin recruitment to damaged mitochondria through phosphorylation of MIC60. Mol. Cell 62, 371–384 (2016).

  214. 214.

    Tsai, P.-I. et al. PINK1 phosphorylates MIC60/mitofilin to control structural plasticity of mitochondrial crista junctions. Mol. Cell 69, 744–756 (2018).

  215. 215.

    Ruan, L. et al. Cytosolic proteostasis through importing of misfolded proteins into mitochondria. Nature 543, 443–446 (2017).

  216. 216.

    Anand, R. et al. The i-AAA protease YME1L and OMA1 cleave OPA1 to balance mitochondrial fusion and fission. J. Cell Biol. 204, 919–929 (2014).

  217. 217.

    Ehses, S. et al. Regulation of OPA1 processing and mitochondrial fusion by m-AAA protease isoenzymes and OMA1. J. Cell Biol. 187, 1023–1036 (2009).

  218. 218.

    Song, Z., Chen, H., Fiket, M., Alexander, C. & Chan, D. C. OPA1 processing controls mitochondrial fusion and is regulated by mRNA splicing, membrane potential, and Yme1L. J. Cell Biol. 178, 749–755 (2007).

  219. 219.

    Mossmann, D. et al. Amyloid-β peptide induces mitochondrial dysfunction by inhibition of preprotein maturation. Cell Metab. 20, 662–669 (2014).

  220. 220.

    Schapira, A. H. Mitochondrial diseases. Lancet 379, 1825–1834 (2012).

  221. 221.

    Okatsu, K., Kimura, M., Oka, T., Tanaka, K. & Matsuda, N. Unconventional PINK1 localization to the outer membrane of depolarized mitochondria drives Parkin recruitment. J. Cell Sci. 128, 964–978 (2015).

  222. 222.

    Bertolin, G. et al. The TOMM machinery is a molecular switch in PINK1 and PARK2/PARKIN-dependent mitochondrial clearance. Autophagy 9, 1801–1817 (2013).

  223. 223.

    Lazarou, M., Jin, S. M., Kane, L. A. & Youle, R. J. Role of PINK1 binding to the TOM complex and alternate intracellular membranes in recruitment and activation of the E3 ligase Parkin. Dev. Cell 22, 320–333 (2012). This study shows that upon dissipation of the mitochondrial membrane potential, the kinase PINK1 accumulates at the mitochondrial outer membrane in a complex with the TOM.

  224. 224.

    Liu, F., Rijkers, D. T. S., Post, H. & Heck, A. J. R. Proteome-wide profiling of protein assemblies by cross-linking mass spectrometry. Nat. Methods 12, 1179–1184 (2015).

  225. 225.

    Huttlin, E. L. et al. Architecture of the human interactome defines protein communities and disease networks. Nature 545, 505–509 (2017).

  226. 226.

    Wai, T. et al. The membrane scaffold SLP2 anchors a proteolytic hub in mitochondria containing PARL and the i-AAA protease YME1L. EMBO Rep. 17, 1844–1856 (2016).

  227. 227.

    Segev, N. & Gerst, J. E. Specialized ribosomes and specific ribosomal protein paralogs control translation of mitochondrial proteins. J. Cell Biol. 217, 117–126 (2018).

  228. 228.

    Hoseini, H. et al. The cytosolic cochaperone Sti1 is relevant for mitochondrial biogenesis and morphology. FEBS J. 283, 3338–3352 (2016).

  229. 229.

    Jores, T. et al. Cytosolic Hsp70 and Hsp40 chaperones enable the biogenesis of mitochondrial β-barrel proteins. J. Cell Biol. 217, 3091–3108 (2018).

  230. 230.

    Opalinski, L. et al. Recruitment of cytosolic J-proteins by TOM receptors promotes mitochondrial protein biogenesis. Cell Rep. 25, 2036–2043 (2018).

  231. 231.

    Hansen, K. G. et al. An ER surface retrieval pathway safeguards the import of mitochondrial membrane proteins in yeast. Science 361, 1118–1122 (2018).

  232. 232.

    Ben-Menachem, R. & Pines, O. Detection of dual targeting and dual function of mitochondrial proteins in yeast. Methods Mol. Biol. 1567, 179–195 (2017).

  233. 233.

    Harsman, A. & Schneider, A. Mitochondrial protein import in trypanosomes: expect the unexpected. Traffic 18, 96–109 (2017).

  234. 234.

    Dienhart, M. K. & Stuart, R. A. The yeast Aac2 protein exists in physical association with the cytochrome bc1-COX supercomplex and the TIM23 machinery. Mol. Biol. Cell 19, 3934–3943 (2008).

  235. 235.

    Takakubo, F. et al. An amino acid substitution in the pyruvate dehydrogenase E1 alpha gene, affecting mitochondrial import of the precursor protein. Am. J. Hum. Genet. 57, 772–780 (1995).

  236. 236.

    Messmer, M. et al. A human pathology-related mutation prevents import of an aminoacyl-tRNA synthetase into mitochondria. Biochem. J. 433, 441–446 (2011).

  237. 237.

    Purdue, P. E., Allsop, J., Isaya, G., Rosenberg, L. E. & Danpure, C. J. Mistargeting of peroxisomal L-alanine:glyoxylate aminotransferase to mitochondria in primary hyperoxaluria patients depends upon activation of a cryptic mitochondrial targeting sequence by a point mutation. Proc. Natl Acad. Sci. USA 88, 10900–10904 (1991).

  238. 238.

    Danpure, C. J., Cooper, P. J., Wise, P. J. & Jennings, P. R. An enzyme trafficking defect in two patients with primary hyperoxaluria type 1: peroxisomal alanine/glyoxylate aminotransferase rerouted to mitochondria. J. Cell Biol. 108, 1345–1352 (1989).

  239. 239.

    Klootwijk, E. D. et al. Mistargeting of peroxisomal EHHADH and inherited renal Fanconi’s syndrome. N. Engl. J. Med. 370, 129–138 (2014).

  240. 240.

    Di Fonzo, A. et al. The mitochondrial disulfide relay system protein GFER is mutated in autosomal-recessive myopathy with cataract and combined respiratory-chain deficiency. Am. J. Hum. Gen. 84, 594–604 (2009).

  241. 241.

    Ceh-Pavia, E., Ang, S. K., Spiller, M. P. & Lu, H. The disease-associated mutation of the mitochondrial thiol oxidase Erv1 impairs cofactor binding during its catalytic reaction. Biochem. J. 464, 449–459 (2014).

  242. 242.

    Shahrour, M. A. et al. Mitochondrial epileptic encephalopathy, 3-methylglutaconic aciduria and variable complex V deficiency associated with TIMM50 mutations. Clin. Genet. 91, 690–696 (2017).

  243. 243.

    Reyes, A. et al. Mutations in TIMM50 compromise cell survival in OxPhos-dependent metabolic conditions. EMBO Mol. Med. 10, e8698 (2018).

  244. 244.

    Ojala, T. et al. New mutation of mitochondrial DNAJC19 causing dilated and noncompaction cardiomyopathy, anemia, ataxia, and male genital anomalies. Pediatr. Res. 72, 432–437 (2012).

  245. 245.

    Davey, K. M. et al. Mutation of DNAJC19, a human homologue of yeast inner mitochondrial membrane co-chaperones, causes DCMA syndrome, a novel autosomal recessive Barth syndrome-like condition. J. Med. Gen. 43, 385–393 (2006).

  246. 246.

    Richter-Dennerlein, R. et al. DNAJC19, a mitochondrial cochaperone associated with cardiomyopathy, forms a complex with prohibitins to regulate cardiolipin remodeling. Cell Metab. 20, 158–171 (2014).

  247. 247.

    Schusdziarra, C., Blamowska, M., Azem, A. & Hell, K. Methylation-controlled J-protein MCJ acts in the import of proteins into human mitochondria. Hum. Mol. Genet. 22, 1348–1357 (2013).

  248. 248.

    Mehawej, C. et al. The impairment of MAGMAS function in human is responsible for a severe skeletal dysplasia. PLOS Genet. 10, e1004311 (2014).

  249. 249.

    Jobling, R. K. et al. PMPCA mutations cause abnormal mitochondrial protein processing in patients with non-progressive cerebellar ataxia. Brain 138, 1505–1517 (2015).

  250. 250.

    Vögtle, F. N. et al. Mutations in PMPCB encoding the catalytic subunit of the mitochondrial presequence protease cause neurodegeneration in early childhood. Am. J. Hum. Gen. 102, 557–573 (2018).

  251. 251.

    Eldomery, M. K. et al. MIPEP recessive variants cause a syndrome of left ventricular non-compaction, hypotonia, and infantile death. Genome Med. 8, 106 (2016).

  252. 252.

    Otto, E. A. et al. Mutation analysis of 18 nephronophthisis associated ciliopathy disease genes using a DNA pooling and next generation sequencing strategy. J. Med. Gen. 48, 105–116 (2011).

  253. 253.

    O’Toole, J. F. et al. Individuals with mutations in XPNPEP3, which encodes a mitochondrial protein, develop a nephronophthisis-like nephropathy. J. Clin. Invest. 120, 791–802 (2010).

  254. 254.

    Magen, D. et al. Mitochondrial Hsp60 chaperonopathy causes an autosomal-recessive neurodegenerative disorder linked to brain hypomyelination and leukodystrophy. Am. J. Hum. Gen. 83, 30–42 (2008).

  255. 255.

    Hansen, J. J. et al. Hereditary spastic paraplegia SPG13 is associated with a mutation in the gene encoding the mitochondrial chaperonin Hsp60. Am. J. Hum. Genet. 70, 1328–1332 (2002).

  256. 256.

    Bie, A. S. et al. Effects of a mutation in the HSPE1 gene encoding the mitochondrial co-chaperonin HSP10 and its potential association with a neurological and developmental disorder. Front. Mol. Biosci. 3, e874 (2016).

  257. 257.

    Koehler, C. M. et al. Human deafness dystonia syndrome is a mitochondrial disease. Proc. Natl Acad. Sci. USA 96, 2141–2146 (1999).

  258. 258.

    Roesch, K., Curran, S. P., Tranebjaerg, L. & Koehler, C. M. Human deafness dystonia syndrome is caused by a defect in assembly of the DDP1/TIMM8a-TIMM13 complex. Hum. Mol. Genet. 11, 477–486 (2002).

  259. 259.

    Mayr, J. A. et al. Lack of the mitochondrial protein acylglycerol kinase causes Sengers syndrome. Am. J. Hum. Gen. 90, 314–320 (2012).

  260. 260.

    Kang, Y. et al. Sengers syndrome-associated mitochondrial acylglycerol kinase is a subunit of the human TIM22 protein import complex. Mol. Cell 67, 457–470 (2017).

  261. 261.

    Vukotic, M. et al. Acylglycerol kinase mutated in sengers syndrome is a subunit of the TIM22 protein translocase in mitochondria. Mol. Cell 67, 471–483 (2017).

  262. 262.

    Kukat, C. et al. Cross-strand binding of TFAM to a single mtDNA molecule forms the mitochondrial nucleoid. Proc. Natl Acad. Sci. USA 112, 11288–11293 (2015).

Download references


This work was supported by European Research Council (ERC) Consolidator Grant No. 648235, the Excellence Initiative and Strategy of the German federal and state governments (EXC 294 BIOSS; GSC-4 Spemann Graduate School; EXC 2189 CIBSS), the Deutsche Forschungsgemeinschaft (PF 202/8-1 and 202/9-1; WA 1598/5-1) and the Sonderforschungsbereiche 746 and 1140.

Author information


  1. Institute of Biochemistry and Molecular Biology, ZBMZ, Faculty of Medicine, University of Freiburg, Freiburg, Germany

    • Nikolaus Pfanner
    •  & Nils Wiedemann
  2. CIBSS Centre for Integrative Biological Signalling Studies, University of Freiburg, Freiburg, Germany

    • Nikolaus Pfanner
    • , Bettina Warscheid
    •  & Nils Wiedemann
  3. Institute of Biology II, Biochemistry – Functional Proteomics, Faculty of Biology, University of Freiburg, Freiburg, Germany

    • Bettina Warscheid


  1. Search for Nikolaus Pfanner in:

  2. Search for Bettina Warscheid in:

  3. Search for Nils Wiedemann in:


The authors contributed equally to all aspects of the article.

Competing interests

The authors declare no competing interests.

Corresponding authors

Correspondence to Nikolaus Pfanner or Nils Wiedemann.


Oxidative phosphorylation

Oxidation of metabolites liberates energy that is used to synthesize ATP; in mitochondria, this is performed by the respiratory chain, which generates a proton gradient across the inner membrane to drive ATP production by the F1F0-ATP synthase.


Membrane-bound machinery that facilitates the insertion of precursor proteins into the lipid phase of a membrane, such as the oxidase assembly (OXA) insertase of the mitochondrial inner membrane.

Mitochondrial contact site and cristae organizing system

(MICOS). A large protein complex of the inner membrane with a dual role: maintenance of the cristae architecture of the inner membrane and the formation of contact sites between the inner and the outer membranes.

TOM complex

The translocase of the outer membrane (TOM) is a protein complex that forms the major mitochondrial entry site for precursor proteins synthesized in the cytosol.

Heat shock protein 70

(Hsp70). A large family of ATP-dependent molecular chaperones of ~70 kDa that bind loosely folded proteins and prevent their misfolding or aggregation. The major mitochondrial heat shock protein 70 (mtHsp70) has a dual role in driving ATP-dependent protein import into the matrix and assisting in folding of proteins.

N-end rule pathway

A pathway in which the amino-terminal amino acid residue links proteins to regulated proteolysis. A destabilizing residue promotes rapid degradation, whereas a stabilizing residue leads to a longer half-life of a protein.

SAM complex

The sorting and assembly machinery (SAM) inserts β-barrel proteins into the mitochondrial outer membrane; it is also known as topogenesis of outer membrane β-barrel proteins (TOB).

Mitochondrial unfolded protein response

(UPRmt). A stress response induced by mitochondrial dysfunction that upregulates the transcription of nuclear genes encoding mitochondrial chaperones, proteases and further components that support mitochondrial recovery and survival.


Large supercomplexes in the mitochondrial inner membrane consisting of complexes I, III and IV of the respiratory chain.

Inner boundary membrane

The mitochondrial inner membrane consists of two domains: the folded cristae, which form invaginations, and the inner boundary membrane, which is located adjacent to the mitochondrial outer membrane.

Crista junctions

Narrow apertures at the beginning of cristae of the mitochondrial inner membrane that link cristae to the inner boundary membrane.

Synthetic growth defects

Mutations or deletions in different genes that result in a stronger growth defect if combined in the same cell.


A mitochondrial DNA–protein assembly located in the matrix containing the packaging factor mitochondrial transcription factor A (TFAM).

ER–mitochondria encounter structure

(ERMES). A multisubunit protein complex that connects the endoplasmic reticulum (ER) and the mitochondrial outer membrane. ERMES is likely involved in lipid transfer between the organelles and is required for maintaining the morphology of mitochondria.

About this article

Publication history


Issue Date