Advertisement

Mitochondria pp 343-360 | Cite as

Studying Proteolysis Within Mitochondria

  • Takashi Tatsuta
  • Thomas Langer
Part of the Methods in Molecular Biology™ book series (MIMB, volume 372)

Abstract

Mitochondria are dynamic organelles with activities that adjust to altering physiological conditions and variable metabolic demands. A conserved proteolytic system present within the organelle exerts essential functions during the biogenesis of mitochondria and ensures the maintenance of organellar activities under varying conditions. Proteases dependent on adenosine triphosphate, in concert with oligopeptidases, degrade nonassembled or damaged proteins in various subcompartments of mitochondria, preventing their accumulation and possibly deleterious effects on mitochondrial functions. Although an increasing number of mitochondrial peptidases are characterized and functionally linked to diverse cellular processes, only limited information is available on the stability of the mitochondrial proteome and the turnover rates of individual proteins. We describe experimental approaches in the yeast Saccharomyces cerevisiae and in mice, allowing analysis of the proteolytic breakdown of mitochondrial proteins individually or on a proteomewide scale.

Key Words

ATP-dependent protease Oligopeptidase Proteolysis quality control 

References

  1. 1.
    Gakh, O., Cavadini, P., and Isaya, G. (2002) Mitochondrial processing peptidases. Biochim. Biophys. Acta 1592, 63–77.PubMedCrossRefGoogle Scholar
  2. 2.
    Esser, K., Tursun, B., Ingenhoven, M., Michaelis, G., and Pratje, E. (2002) A novel two-step mechanism for removal of a mitochondrial signal sequence involves the m-AAA complex and the putative rhomboid protease Pcp1. J. Mol. Biol. 323, 835–843.PubMedCrossRefGoogle Scholar
  3. 3.
    Herlan, M., Vogel, F., Bornhövd, C., Neupert, W., and Reichert, A.S. (2003) Processing of Mgm1 by the rhomboid-type protease Pcp1 is required for maintenance of mitochondrial morphology and of mitochondrial DNA. J. Biol. Chem. 278, 27,781–27,788.PubMedCrossRefGoogle Scholar
  4. 4.
    McQuibban, G. A., Saurya, S., and Freeman, M. (2003) Mitochondrial membrane remodelling regulated by a conserved rhomboid protease. Nature 423, 537–541.PubMedCrossRefGoogle Scholar
  5. 5.
    Van Dyck, L. and Langer, T. (1999) ATP-dependent proteases controlling mitochondrial function in the yeast Saccharomyces cerevisiae. Cell Mol. Life Sci. 55, 825–842.CrossRefGoogle Scholar
  6. 6.
    Bota, D. A. and Davies, K. J. A. (2001) Protein degradation in mitochondria: implications for oxidative stress, aging and disease: a novel etiological classification of mitochondrial proteolytic disorders. Mitochondrion 1, 33–49.PubMedCrossRefGoogle Scholar
  7. 7.
    Young, L., Leonhard, K., Tatsuta, T., Trowsdale, J., and Langer, T. (2001) Role of the ABC transporter Mdl1 in peptide export from mitochondria. Science 291, 2135–2138.PubMedCrossRefGoogle Scholar
  8. 8.
    Augustin, S., Nolden, M., Müller, S., Hardt, O., Arnold, I., and Langer, T. (2005) Characterization of peptides released from mitochondria: evidence for constant proteolysis and peptide efflux. J. Biol. Chem. 280, 2691–2699.PubMedCrossRefGoogle Scholar
  9. 9.
    Kambacheld, M., Augustin, S., Tatsuta, T., Müller, S., and Langer, T. (2005) Role of the novel metallopeptidase MOP112 and saccharolysin for the complete degradation of proteins residing in different subcompartments of mitochondria. J. Biol. Chem. 280, 20,132–20,139.PubMedCrossRefGoogle Scholar
  10. 10.
    Daum, G., Gasser, S. M., and Schatz, G. (1982) Import of proteins into mitochondria. Energy-dependent, two-step processing of the intermembrane space enzyme cytochrome b2 by isolated yeast mitochondria. J. Biol. Chem. 257, 13,075–13,080.PubMedGoogle Scholar
  11. 11.
    Herrmann, J. M., Fölsch, H., Neupert, W., and Stuart, R. A. (1994) Isolation of yeast mitochondria and study of mitochondrial protein translation, in Cell Biology: A Laboratory Handbook, vol. 1 (Celis, D. E., ed.), Academic Press, San Diego, CA, pp. 538–544.Google Scholar
  12. 12.
    Meisinger, C., Sommer, T., and Pfanner, N. (2000) Purification of Saccharomcyes cerevisiae mitochondria devoid of microsomal and cytosolic contaminations. Anal. Biochem. 287, 339–342.PubMedCrossRefGoogle Scholar
  13. 13.
    Brandt, A. (1991) Pulse labeling of yeast cells as a tool to study mitochondrial protein import. Methods Cell Biol. 34, 369–376.PubMedCrossRefGoogle Scholar
  14. 14.
    Kozak, M. (1987) An analysis of 5′-noncoding sequences from 699 vertebrate messenger RNAs. Nucleic Acids Res. 15, 8125–8148.PubMedCrossRefGoogle Scholar
  15. 15.
    Black-Schaefer, C. L., McCourt, J. D., Poyton, R. O., and McKee, E. E. (1991) Mitochondrial gene expression in Saccharomyces cerevisiae. Proteolysis of nascent chains in isolated yeast mitochondria optimized for protein synthesis. Biochem. J. 274, 199–205.PubMedGoogle Scholar
  16. 16.
    Mattiazzi, M., D’Aurelio, M., Gajewski, C. D., et al. (2002) Mutated human SOD1 causes dysfunction of oxidative phosphorylation in mitochondria of transgenic mice. J. Biol. Chem. 277, 29,626–29,633.PubMedCrossRefGoogle Scholar
  17. 17.
    Gancedo, J. M. (1998) Yeast carbon catabolite repression. Microbiol. Mol. Biol. Rev. 62, 334–361.PubMedGoogle Scholar
  18. 18.
    Käser, M., Kambacheld, M., Kisters-Woike, B., and Langer, T. (2003) Oma1, a novel membrane-bound metallopeptidase in mitochondria with activities overlapping with the m-AAA protease. J. Biol. Chem. 278, 46,414–46,423.PubMedCrossRefGoogle Scholar
  19. 19.
    Leonhard, K., Guiard, B., Pellechia, G., Tzagoloff, A., Neupert, W., and Langer, T. (2000) Membrane protein degradation by AAA proteases in mitochondria: extraction of substrates from either membrane surface. Mol. Cell 5, 629–638.PubMedCrossRefGoogle Scholar
  20. 20.
    Rottgers, K., Zufall, N., Guiard, B., and Voos, W. (2002) The ClpB homolog Hsp78 is required for the efficient degradation of proteins in the mitochondrial matrix. J. Biol. Chem. 277, 45,829–45,837.PubMedCrossRefGoogle Scholar
  21. 21.
    Leonhard, K., Stiegler, A., Neupert, W., and Langer, T. (1999) Chaperone-like activity of the AAA domain of the yeast Yme1 AAA protease. Nature 398, 348–351.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 2007

Authors and Affiliations

  • Takashi Tatsuta
    • 1
  • Thomas Langer
    • 1
  1. 1.Institut für GenetikUniversität zu KölnKölnGermany

Personalised recommendations