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Reconstitution reveals Ykt6 as the autophagosomal SNARE in autophagosome–vacuole fusion

Bas, Levent, Papinski, Daniel, Licheva, Mariya, Torggler, Raffaela, Rohringer, Sabrina, Schuschnig, Martina, Kraft, Claudine·2018·Journal of Cell Biology
DOI 10.1083/jcb.201804028PMC PMC6168255
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Abstract

Autophagy mediates the bulk degradation of cytoplasmic material, particularly during starvation. Upon the induction of autophagy, autophagosomes form a sealed membrane around cargo, fuse with a lytic compartment, and release the cargo for degradation. The mechanism of autophagosome–vacuole fusion is poorly understood, although factors that mediate other cellular fusion events have been implicated. In this study, we developed an in vitro reconstitution assay that enables systematic discovery and dissection of the players involved in autophagosome–vacuole fusion. We found that this process requires the Atg14–Vps34 complex to generate PI3P and thus recruit the Ypt7 module to autophagosomes. The HOPS-tethering complex, recruited by Ypt7, is required to prepare SNARE proteins for fusion. Furthermore, we discovered that fusion requires the R-SNARE Ykt6 on the autophagosome, together with the Q-SNAREs Vam3, Vam7, and Vti1 on the vacuole. These findings shed new light on the mechanism of autophagosome–vacuole fusion and reveal that the R-SNARE Ykt6 is required for this process.

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  • Both autophagosome-vacuole fusion in yeast and autophagosome-lysosome fusion in mammals have been recently reconstituted using partially purified fractions of autophagosomes, vacuoles/lysosomes and cytosol.

    “Both autophagosome-vacuole fusion in yeast and autophagosome-lysosome fusion in mammals has been recently reconstituted, using partially purified fractions of autophagosomes, vacuoles/lysosomes and cytosol [506-508].”

    From Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition) 1 (2021) · cites this paper as [506]

  • In vitro reconstitutions of autophagosome-vacuole and autophagosome-lysosome fusion have partially overcome the problem of analyzing fusion in intact cells and made it possible to identify relevant proteins and their functions in this specific step of autophagy.

    “In vitro reconstitutions of autophagosome-vacuole and autophagosome-lysosome fusion have partially overcome this problem and made it possible to identify relevant proteins and their functions in this specific step of autophagy.”

    From Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition) 1 (2021) · cites this paper as [506]

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Metadata

Authors
Bas, L., Papinski, D., Licheva, M., Torggler, R., Rohringer, S., Schuschnig, M., & Kraft, C.
Affiliations
  • Max F. Perutz Laboratories, Vienna Biocenter, University of Vienna, Vienna, Austria 1
  • Institute of Biochemistry and Molecular Biology, Centre for Biochemistry and Molecular Cell Research , Faculty of Medicine, University of Freiburg, Freiburg, Germany 2
ORCID
Venue
Journal of Cell Biology
Publisher
Rockefeller University Press
Published
2018-08-10
Volume
217
Issue
10
Pages
3656-3669
ISSN
  • 0021-9525
  • 1540-8140
DOI
10.1083/jcb.201804028
PMC
PMC6168255
Cited by
111
References
2
Language
en
Licence
creativecommons.org/licenses/by/4.0/ (vor, 467 d)
Funders
  • Vienna Science and Technology Fund (VRG10-001)
  • Austrian Science Fund (P25522-B20, P28113-B28)
  • European Research Council (769065, 765912)
  • European Molecular Biology Organization
  • Österreichische Akademie der Wissenschaften

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