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.

  • Review Article
  • Published:

Ubiquitin and proteasomes

Molecular dissection of autophagy: two ubiquitin-like systems

Nature Reviews Molecular Cell Biology volume 2, pages 211–216 (2001)Cite this article

Key Points

  • This review discusses recent analyses of the genes required for autophagy ? intracellular bulk protein degradation ? in yeast, where two ubiquitin-like systems have now been revealed.

  • Light microscopy studies show that starved yeast cells take up their own cytoplasm into vacuoles through autophagic bodies. Closer analysis using electron microscopy show that these bodies form a double-membraned structure called the autophagosome, which subsequently fuses with the vacuole/lysosome. The whole process is topologically the same in mammals.

  • Screens in yeast defective in autophagy morphologically and biochemically revealed two sets of genes, the APG and AUT genes, respectively. A specific vacuolar enzyme biosynthetic pathway requires the cytosol-to-vacuole-targeting (CVT) genes. There is extensive overlap between the CVT genes and the APG/AUT genes.All apg mutants have defects at or before formation of the autophagosome.

  • Two ubiquitin-like systems have been discovered. The first ? the Apg12 conjugation system ? is unique in that Apg12 is synthesized as a mature form; it seems to have just one target, Apg5, and at steady state almost all Apg12 molecules are conjugated with Apg5.

  • The second ? Apg8/Aut7 ? is processed at its carboxy-terminal region by Aut2/Apg4. Apg8 exists in two forms, one of which is membrane bound through a phospholipid. This lipidation is mediated by ubiqutin-like system; Apg8 is activated by Apg7 and transferred to Apg3 and finally forms a conjugate with phosphatidylethaolamine (PE). Apg4 cleaves Apg8?PE, releasing Apg8 from membrane.

  • Morphological studies show that Apg8 localizes on the membrane of intermediate structures of the autophagosome; this transient association seems to be essential for formation of the autophagosome.

  • Both Apg12 and Apg8 are highly conserved, with apparent homologues in the worm, mammals and plants. In higher eukaryotes, Apg8 consists of a multigene family.

Abstract

Recent analyses of the genes required for autophagy ? intracellular bulk protein degradation ? in yeast have revealed two ubiquitin-like systems, both of which are involved in the membrane dynamics of the process. Molecular dissection of these systems is now revealing some surprises.

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: Electron microscopic images of autophagy in yeast.
Figure 2: Two ubiquitin-like systems are required for autophagy.

Similar content being viewed by others

Notes

  1. *Macroautophagy is generally a non-selective sequestration.But in some physiological situations, excess or unnecessary organelles areselectively degraded in a lysosome/vacuole. The best-studied case is peroxisomedegradation, called pexophagy by Daniel Klionsky. In the yeast Pichia pastoris, peroxisomes are sequestered through either macroautophagy or microautophagy,depending on the nutrient conditions40.

References

  1. Hershko, A. & Ciechanover, A. The ubiquitin system. Annu. Rev. Biochem. 67, 425?479 (1998).

    Article  CAS  Google Scholar 

  2. Klionsky, D. J. & Ohsumi, Y. Vacuolar import of proteins and organelles from the cytoplasm. Annu. Rev. Cell Dev. Biol. 15, 1?32 ( 1999).

    Article  CAS  Google Scholar 

  3. Baba, M., Takeshige, K., Baba, N. & Ohsumi, Y. Ultrastructural analysis of the autophagic process in yeast: detection of autophagosomes and their characterization. J. Cell Biol. 124, 903?913 (1994).

    Article  CAS  Google Scholar 

  4. Takeshige, K., Baba, M., Tsuboi, S., Noda, T. & Ohsumi, Y. Autophagy in yeast demonstrated with proteinase-deficient mutants and conditions for its induction. J. Cell Biol. 119, 301?311 (1992). This is the first report that yeast induces autophagy that is quite similar to that in mammals under various starvation conditions.

    Article  CAS  Google Scholar 

  5. Kim, J. & Klionsky, D. J. Autophagy, the cytoplasm-to-vacuole-targeting pathway, and pexophagy in yeast and mammalian cells. Annu. Rev. Biochem. 69, 303?342 ( 2000).

    Article  CAS  Google Scholar 

  6. Klionsky, D. J. & Emr, S. Autophagy as a regulated pathway of cellular degradation. Science 290, 1717?1721 (2000).

    Article  CAS  Google Scholar 

  7. Baba, M., Osumi, M. & Ohsumi, Y. Analysis of the membrane structures involved in autophagy in yeast by freeze-replica method. Cell Struct. Funct. 20, 465?471 (1995).

    Article  CAS  Google Scholar 

  8. Tsukada, M. & Ohsumi, Y. Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae. FEBS Lett. 333, 169?174 (1993).

    Article  CAS  Google Scholar 

  9. Thumm, M. et al. Isolation of autophagocytosis mutants of Saccharomyces cerevisiae . FEBS Lett. 349, 275? 280 (1994).

    Article  CAS  Google Scholar 

  10. Mizushima, N., Noda, T. & Ohsumi, Y. Apg16p is required for the function of the Apg12p?Apg5p conjugate in the yeast autophagy pathway. EMBO J. 18 , 3888?3896 (1999).

    Article  CAS  Google Scholar 

  11. Kamada, Y. et al. Tor-mediated induction of autophagy via an Apg1 protein kinase complex. J. Cell Biol. 150, 1507? 1513 (2000).

    Article  CAS  Google Scholar 

  12. Harding, T. M., Morano, K. A., Scott, S. V. & Klionsky, D. J. Isolation and characterization of yeast mutants in the cytoplasm to vacuole protein targeting pathway. J. Cell Biol. 131, 591?602 (1995).

    Article  CAS  Google Scholar 

  13. Noda, T. et al. Apg9p/Cvt7p is an integral membrane protein required for transport vesicle formation in the Cvt and autophagy pathways. J. Cell Biol. 148, 465?480 ( 2000).

    Article  CAS  Google Scholar 

  14. George, M. D. et al. Apg5p functions in the sequestration step in the cytoplasm-to-vacuole targeting and macroautophagy pathways. Mol. Biol. Cell 11, 969?982 (2000).

    Article  CAS  Google Scholar 

  15. Yuan, W., Stromhaug, P. E. & Dunn, W. A. Jr Glucose-induced autophagy of peroxisomes in Pichia pastori requires a unique E1-like protein. Mol. Biol. Cell 10, 1353?1366 (1999).

    Article  CAS  Google Scholar 

  16. Kim, J., Dalton, V. M., Eggerton, K. P., Scott, S. V. & Klionsky, D. J. Apg7p/Cvt2p is required for the cytoplasm-to-vacuole targeting, macroautophagy, and peroxisome degradation pathways. Mol. Biol. Cell 10, 1337? 1351 (1999).

    Article  CAS  Google Scholar 

  17. Hutchins, M. U., Veenhuis, M. & Klionsky, D. J. Peroxisome degradation in Saccharomyces cerevisiae is dependent on machinery of macroautophagy and the Cvt pathway. J. Cell Sci. 112, 4079?4087 (1999).

    CAS  PubMed  Google Scholar 

  18. Muller, O. et al. Autophagic tubes. Vacuolar invaginations involved in lateral membrane sorting and inverse vesicle budding. J. Cell Biol. 151, 519?528 (2000).

    Article  CAS  Google Scholar 

  19. Suriapranata, I. et al. The breakdown of autophagic vesicles inside the vacuole depends on Aut4p. J. Cell Sci. 113, 4025? 4033 (2000).

    CAS  PubMed  Google Scholar 

  20. Mizushima, N. et al. A protein conjugation system essential for autophagy. Nature 395, 395?398 ( 1998).This paper presented the first ubiquitin-like protein conjugation reaction, the Apg12 system, as being essential for autophagy.

    Article  CAS  Google Scholar 

  21. Tanida, I. et al. Apg7p/Cvt2p: A novel protein-activating enzyme essential for autophagy. Mol. Biol. Cell 10, 1367? 1379 (1999).

    Article  CAS  Google Scholar 

  22. Shintani, T. et al. Apg10p, a novel protein-conjugating enzyme essential for autophagy in yeast. EMBO J. 18, 5234? 5241 (1999).

    Article  CAS  Google Scholar 

  23. Kirisako, T. et al. Formation process of autophagosome is traced with Apg8/Aut7p in yeast. J. Cell Biol. 147, 435? 446 (1999).

    Article  CAS  Google Scholar 

  24. Huang, W. P., Scott, S. V., Kim, J. & Klionsky, D. J. The itinerary of a vesicle component, Aut7p/Cvt5p, terminates in the yeast vacuole via the autophagy/Cvt pathways. J. Biol. Chem. 275, 5845?5851 (2000).

    Article  CAS  Google Scholar 

  25. Lang, T. et al. Aut2p and Aut7p, two novel microtubule-associated proteins are essential for delivery of autophagic vesicles to the vacuole. EMBO J. 17, 3597?3607 ( 1998).

    Article  CAS  Google Scholar 

  26. Kirisako, T. et al. The reversible modification regulates the membrane-binding state of Apg8/Aut7 essential for autophagy and the cytoplasm to vacuole targeting pathway. J. Cell Biol. 151, 263? 276 (2000).This showed that Apg8 associates with the intermediate membranes of the autophagosome by serial modification reactions.

    Article  CAS  Google Scholar 

  27. Ichimura, Y. et al. A ubiquitin-like system mediates protein lipidation. Nature 408, 489?493 ( 2000).This paper described the Apg8 lipidation system, in which the processed form of Apg8 is activated by a ubiquitin-like system and finally forms a conjugate with the membrane phospholipid, phosphatidylethanolamine. PubMed

    Article  Google Scholar 

  28. Mizushima, N., Sugita, H., Yoshimori, T. & Ohsumi, Y. A new protein conjugation system in human. The counterpart of the yeast Apg12p conjugation system essential for autophagy. J. Biol. Chem. 273, 33889?33892 (1998).

    Article  CAS  Google Scholar 

  29. Mizushima, N. et al. Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J.Cell Biol. (in the press).

  30. Mann, S. S. & Hammarback, J. A. Molecular characterization of light chain 3. A microtubule binding subunit of MAP1A and MAP1B. J. Biol. Chem. 269, 11492?11497 (1994).

    CAS  PubMed  Google Scholar 

  31. Kabeya, Y. et al. LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. EMBO J. 19 , 5720?5728 (2000). Two UBL systems, Apg12 and Apg8, are conserved in higher eukaryotes and this is the first report of the functional homologue of Apg8 in mammals.

    Article  CAS  Google Scholar 

  32. Meyers, G., Stoll, D. & Gunn, M. Insertion of a sequence encoding light chain 3 of microtubule-associated proteins 1A and 1B in a pestivirus genome: connection with virus cytopathogenicity and induction of lethal disease in cattle. J. Virol. 72, 4139?4148 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Sagiv, Y., Legesse-Miller, A., Porat, A. & Elazar, Z. GATE-16, a membrane transport modulator, interacts with NSF and the Golgi v-SNARE GOS-28. EMBO J. 19, 1494? 1504 (2000).

    Article  CAS  Google Scholar 

  34. Legesse-Miller, A., Sagiv, Y., Glozman, R. & Elazar, Z. Aut7p, a soluble autophagic factor, participates in multiple membrane trafficking processes . J. Biol. Chem. 275, 32966? 32973 (2000).

    Article  CAS  Google Scholar 

  35. Chen, L., Wang, H., Vicini, S. & Olsen, R. W. The γ-aminobutyric acid type A (GABAA) receptor-associated protein (GABARAP) promotes GABAA receptor clustering and modulates the channel kinetics. Proc. Natl Acad. Sci. USA 97, 11557? 11562 (2000).

    Article  CAS  Google Scholar 

  36. Wang, H., Bedford, F. K., Brandon, N. J., Moss, S. J. & Olsen, R. W. GABAA-receptor-associated protein links GABAA receptors and the cytoskeleton. Nature 397, 69?72 ( 1999).

    Article  CAS  Google Scholar 

  37. Paz, Y., Elazar, Z. & Fass, D. Structure of GATE-16, membrane transport modulator and mammalian ortholog of autophagocytosis factor Aut7p. J. Biol. Chem. 275, 25445?25450 ( 2000).

    Article  CAS  Google Scholar 

  38. Suzuki, K., Kirisako, T. & Ohsumi, Y. Localization of Apg5p and Apg8p/Aut7p reveals functional classes of APG genes and their interactions in autophagy. (submitted).

  39. Wendland, B., Emr, S. D. & Riezman, H. Protein traffic in the yeast endocytic and vacuolar protein sorting pathways. Curr. Opin. Cell Biol. 10 , 513?522 (1998).

    Article  CAS  Google Scholar 

  40. Tuttle, D. L. & Dunn, W. A. Divergent modes of autophagy in the methylotrophic yeast Pichia pastoris. J. Cell Sci. 108, 25?35 (1995).

    CAS  PubMed  Google Scholar 

  41. Huang, P. H. & Chiang, H. L. Identification of novel vesicles in the cytosol to vacuole protein degradation pathway. J. Cell Biol. 136, 803?810 ( 1997).

    Article  CAS  Google Scholar 

  42. Cuervo, A. M. & Dice, J. F. Age-related decline in chaperone-mediated autophagy. J. Biol. Chem. 275, 31505? 31513 (2000).

    Article  CAS  Google Scholar 

  43. Scott, S. V., Baba, M., Ohsumi, Y. & Klionsky, D. J. Aminopeptidase I is targeted to the vacuole by a nonclassical vesicular mechanism. J. Cell Biol. 138, 37?44 (1997).

    Article  CAS  Google Scholar 

  44. Baba, M., Osumi, M., Scott, S. V., Klionsky, D. J. & Ohsumi, Y. Two distinct pathways for targeting proteins from the cytoplasm to the vacuole/lysosome. J. Cell Biol. 139 , 1687?1695 (1997).

    Article  CAS  Google Scholar 

  45. Harding, T. M., Hefner-Gravink, A., Thumm, M. & Klionsky, D. J. Genetic and phenotypic overlap between autophagy and the cytoplasm to vacuole protein targeting pathway. J. Biol. Chem. 271, 17621?17624 (1996).

    Article  CAS  Google Scholar 

  46. Scott, S. V. et al. Cytoplasm-to-vacuole targeting and autophagy employ the same machinery to deliver proteins to the yeast vacuole. Proc. Natl Acad. Sci. USA 93, 12304?12308 (1996).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

Thanks to T. Noda and N. Mizushima for critical discussion and reading of this manuscript.

Author information

Authors and Affiliations

  1. Department of Cell Biology, National Institute for Basic Biology, Myodaiji, 444-8585 , Okazaki, Japan

    Yoshinori Ohsumi

Authors
  1. Yoshinori Ohsumi
    View author publications

    You can also search for this author in PubMed Google Scholar

Related links

Related links

DATABASE LINKS

APG16

APG17

Apg12

Apg1

apg6

vps30

Apg7/Cvt2

Apg10

Aut7/Apg8/Cvt5

Aut2/Apg4

Aut1

Apg3

Apg5

MAP1-LC3

GABARAP

ENCYCLOPEDIA OF LIFE SCIENCES

Lysosomal degradation of proteins

Glossary

E1 ENZYME

An enzyme that activates the carboxy-terminal glycine of the small protein ubiquitin, or ubiquitin-like proteins, allowing them to form a high-energy bond to a specific cysteine residue of the E1.

E2 ENZYME

An enzyme that accepts ubiquitin or a ubiquitin-like protein from an E1 and transfers it to the substrate, mostly using an E3 enzyme.

NSF

N-ethylmaleimide-sensitive factor, an AAA-type ATPase essential for membrane fusion during vesicle transport.

V-SNARE

A family of proteins on secretory vesicles. They form a complex with t-SNAREs on the target membrane during vesicle fusion.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ohsumi, Y. Molecular dissection of autophagy: two ubiquitin-like systems. Nat Rev Mol Cell Biol 2, 211–216 (2001). https://doi.org/10.1038/35056522

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1038/35056522

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing