Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Stendomycin selectively inhibits TIM23-dependent mitochondrial protein import

Nature Chemical Biology volume 13, pages 1239–1244 (2017)Cite this article

Subjects

Abstract

Tim17 and Tim23 are the main subunits of the TIM23 complex, one of the two major essential mitochondrial inner-membrane protein translocon machineries (TIMs). No chemical probes that specifically inhibit TIM23-dependent protein import were known to exist. Here we show that the natural product stendomycin, produced by Streptomyces hygroscopicus, is a potent and specific inhibitor of the TIM23 complex in yeast and mammalian cells. Furthermore, stendomycin-mediated blockage of the TIM23 complex does not alter normal processing of the major regulatory mitophagy kinase PINK1, but TIM23 is required to stabilize PINK1 on the outside of mitochondria to initiate mitophagy upon membrane depolarization.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Stendomycin targets the Tim17–Tim23 complex.
Figure 2: Stendomycin blocks TIM23-mediated, but not TIM22-mediated, mitochondrial protein import.
Figure 3: Stendomycin inhibits TIM23-dependent translocation in vivo.
Figure 4: Mechanism of action of stendomycin is conserved.
Figure 5: Stendomycin inhibits mitochondrial inner-membrane translocation but does not stabilize PINK1 or trigger mitophagy.

Similar content being viewed by others

References

  1. Aoun, M. & Tiranti, V. Mitochondria: a crossroads for lipid metabolism defect in neurodegeneration with brain iron accumulation diseases. Int. J. Biochem. Cell Biol. 63, 25–31 (2015).

    Article  CAS  Google Scholar 

  2. Bhola, P.D. & Letai, A. Mitochondria-judges and executioners of cell death Sentences. Mol. Cell 61, 695–704 (2016).

    Article  CAS  Google Scholar 

  3. Endo, T., Yamamoto, H. & Esaki, M. Functional cooperation and separation of translocators in protein import into mitochondria, the double-membrane bounded organelles. J. Cell Sci. 116, 3259–3267 (2003).

    Article  CAS  Google Scholar 

  4. Chacinska, A., Koehler, C.M., Milenkovic, D., Lithgow, T. & Pfanner, N. Importing mitochondrial proteins: machineries and mechanisms. Cell 138, 628–644 (2009).

    Article  CAS  Google Scholar 

  5. MacKenzie, J.A. & Payne, R.M. Mitochondrial protein import and human health and disease. Biochim. Biophys. Acta. 1772, 509–523 (2007).

    Article  CAS  Google Scholar 

  6. Pfanner, N. & Meijer, M. Mitochondrial biogenesis: the Tom and Tim machine. Curr. Biol. 7, R100–R103 (1997).

    Article  CAS  Google Scholar 

  7. Thompson, R.Q. & Hughes, M.S. Stendomycin: A new antifungal antibiotic. J. Antibiot. (Tokyo) 16, 187–194 (1963).

    CAS  Google Scholar 

  8. Hoepfner, D. et al. High-resolution chemical dissection of a model eukaryote reveals targets, pathways and gene functions. Microbiol. Res. 169, 107–120 (2014).

    Article  CAS  Google Scholar 

  9. Saleem, A., Iqbal, S., Zhang, Y. & Hood, D.A. Effect of p53 on mitochondrial morphology, import, and assembly in skeletal muscle. Am. J. Physiol. Cell Physiol. 308, C319–C329 (2015).

    Article  CAS  Google Scholar 

  10. Mokranjac, D. & Neupert, W. The many faces of the mitochondrial TIM23 complex. Biochim. Biophys. Acta. 1797, 1045–1054 (2010).

    Article  CAS  Google Scholar 

  11. 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).

    Article  CAS  Google Scholar 

  12. Jensen, R.E. & Johnson, A.E. Protein translocation: is Hsp70 pulling my chain? Curr. Biol. 9, R779–R782 (1999).

    Article  CAS  Google Scholar 

  13. 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).

    Article  Google Scholar 

  14. Chacinska, A. et al. Mitochondrial presequence translocase: switching between TOM tethering and motor recruitment involves Tim21 and Tim17. Cell 120, 817–829 (2005).

    Article  CAS  Google Scholar 

  15. Greene, A.W. et al. Mitochondrial processing peptidase regulates PINK1 processing, import and Parkin recruitment. EMBO Rep. 13, 378–385 (2012).

    Article  CAS  Google Scholar 

  16. Rüb, C., Wilkening, A. & Voos, W. Mitochondrial quality control by the Pink1/Parkin system. Cell Tissue Res. 367, 111–123 (2017).

    Article  Google Scholar 

  17. Becker, D., Richter, J., Tocilescu, M.A., Przedborski, S. & Voos, W. Pink1 kinase and its membrane potential (ΔΨ)-dependent cleavage product both localize to outer mitochondrial membrane by unique targeting mode. J. Biol. Chem. 287, 22969–22987 (2012).

    Article  CAS  Google Scholar 

  18. 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).

    Article  CAS  Google Scholar 

  19. Hasson, S.A. et al. High-content genome-wide RNAi screens identify regulators of parkin upstream of mitophagy. Nature 504, 291–295 (2013).

    Article  CAS  Google Scholar 

  20. Kersten, R.D. et al. A mass spectrometry-guided genome mining approach for natural product peptidogenomics. Nat. Chem. Biol. 7, 794–802 (2011).

    Article  CAS  Google Scholar 

  21. Ungermann, C., Guiard, B., Neupert, W. & Cyr, D.M. The delta psi- and Hsp70/MIM44-dependent reaction cycle driving early steps of protein import into mitochondria. EMBO J. 15, 735–744 (1996).

    Article  CAS  Google Scholar 

  22. Schneider, H.-C. et al. Mitochondrial Hsp70/MIM44 complex facilitates protein import. Nature 371, 768–774 (1994).

    Article  CAS  Google Scholar 

  23. Simorre, J.P., Genest, D., Caille, A. & Ptak, M. A 2D NMR study of the internal flexibility of the antifungal peptide stendomycin. Eur. Biophys. J. 18, 309–316 (1990).

    Article  CAS  Google Scholar 

  24. Huang, Z. et al. A functional variomics tool for discovering drug-resistance genes and drug targets. Cell Rep. 3, 577–585 (2013).

    Article  CAS  Google Scholar 

  25. Pries, V. et al. Advantages and challenges of phenotypic screens: the identification of two novel antifungal geranylgeranyltransferase I inhibitors. J. Biomol. Screen. 21, 306–315 (2016).

    Article  CAS  Google Scholar 

  26. Glick, B.S. & Pon, L.A. Isolation of highly purified mitochondria from Saccharomyces cerevisiae. Methods Enzymol. 260, 213–223 (1995).

    Article  CAS  Google Scholar 

  27. Claypool, S.M., Whited, K., Srijumnong, S., Han, X. & Koehler, C.M. Barth syndrome mutations that cause tafazzin complex lability. J. Cell Biol. 192, 447–462 (2011).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors acknowledge the NIH GM61721 to C.M.K. and DFG STE 2045/1-1 to J.S. The authors also wish to thank the rest of the Novartis HIP HOP team.

Author information

Author notes

  1. Raffaele Cerino

    Present address: Pharma Research and Early Development, Roche, Basel, Switzerland

  2. Ireos Filipuzzi and Janos Steffen: These authors contributed equally to this work.

Authors and Affiliations

  1. Novartis Institutes for BioMedical Research, Basel, Switzerland

    Ireos Filipuzzi, Mitchel Germain, Laetitia Goepfert, Christoph Potting, Raffaele Cerino, Martin Pfeifer, Philipp Krastel, Dominic Hoepfner, Julie Bastien & Stephen B Helliwell

  2. Department of Chemistry and Biochemistry, University of California, Los Angeles, Los Angeles, California, USA

    Janos Steffen, Michael A Conti & Carla M Koehler

Authors
  1. Ireos Filipuzzi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Janos Steffen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mitchel Germain
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Laetitia Goepfert
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Michael A Conti
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Christoph Potting
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Raffaele Cerino
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Martin Pfeifer
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Philipp Krastel
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Dominic Hoepfner
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Julie Bastien
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Carla M Koehler
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Stephen B Helliwell
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

I.F. and J.S. designed and performed experiments, analyzed the data and wrote the paper; M.G., L.G., M.A.C., C.P., R.C. and M.P., designed and performed experiments and analyzed the data; P.K. and D.H. supervised experiments and analyzed the data; J.B. designed experiments and analyzed the data; C.M.K. and S.B.H. designed experiments, analyzed the data and wrote the paper.

Corresponding authors

Correspondence to Carla M Koehler or Stephen B Helliwell.

Ethics declarations

Competing interests

I.F., M.G., L.G., C.P., R.C., M.P., P.K., D.H., J.B. and S.B.H. are current or former employees of Novartis and may hold stock in the company.

Supplementary information

Supplementary Text and Figures

Supplementary Results, Supplementary Figures 1–5 and Supplementary Note. (PDF 22313 kb)

Reporting Summary (PDF 129 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Filipuzzi, I., Steffen, J., Germain, M. et al. Stendomycin selectively inhibits TIM23-dependent mitochondrial protein import. Nat Chem Biol 13, 1239–1244 (2017). https://doi.org/10.1038/nchembio.2493

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.2493

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research