Skip to main content

Thank you for visiting 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.

A subdomain of the endoplasmic reticulum forms a cradle for autophagosome formation


Autophagy is a bulk degradation process in eukaryotic cells and has fundamental roles in cellular homeostasis.The origin and source of autophagosomal membranes are long-standing questions in the field. Using electron microscopy, we show that, in mammalian culture cells, the endoplasmic reticulum (ER) associates with early autophagic structures called isolation membranes (IMs). Overexpression of an Atg4B mutant, which causes defects in autophagosome formation, induces the accumulation of ER–IM complexes. Electron tomography revealed that the ER–IM complex appears as a subdomain of the ER that formed a cradle encircling the IM, and showed that both ER and isolation membranes are interconnected.

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

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Get just this article for as long as you need it


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

Figure 1: Close association of the ER with the IM during autophagy.
Figure 2: Immunogold electron microscopy of ER–IM complexes.
Figure 3: Electron tomography of ER–IM complexes from control cells.
Figure 4: Electron tomography of two different ER–IM complexes shows a connection between the IM and the ER.
Figure 5: Possible models for autophagosome formation in mammalian cells.


  1. Mizushima, N. Autophagy: process and function. Genes Dev. 21, 2861–2873 (2007).

    CAS  PubMed  Google Scholar 

  2. Suzuki, K. & Ohsumi, Y. Molecular machinery of autophagosome formation in yeast, Saccharomyces cerevisiae. FEBS Lett. 581, 2156–2161 (2007).

    Article  CAS  PubMed  Google Scholar 

  3. Yoshimori, T. & Noda, T. Toward unraveling membrane biogenesis in mammalian autophagy. Curr. Opin. Cell Biol. 20, 401–407 (2008).

    Article  CAS  PubMed  Google Scholar 

  4. Juhasz, G. & Neufeld, T. P. Autophagy: a forty-year search for a missing membrane source. PLoS Biol. 4, e36 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  5. Dunn, W.A. Jr Autophagy and related mechanisms of lysosome-mediated protein degradation. Trends Cell Biol. 4, 139–143 (1994).

    Article  CAS  PubMed  Google Scholar 

  6. Furuno, K. et al. Immunocytochemical study of the surrounding envelope of autophagic vacuoles in cultured rat hepatocytes. Exp. Cell Res. 189, 261–268 (1990).

    Article  CAS  PubMed  Google Scholar 

  7. Yamamoto, A., Masaki, R., Fukui, Y. & Tashiro, Y. Absence of cytochrome P-450 and presence of autolysosomal membrane antigens on the isolation membranes and autophagosomal membranes in rat hepatocytes. J. Histochem. Cytochem. 38, 1571–1581 (1990).

    Article  CAS  PubMed  Google Scholar 

  8. Yamamoto, A., Masaki, R. & Tashiro, Y. Characterization of the isolation membranes and the limiting membranes of autophagosomes in rat hepatocytes by lectin cytochemistry. J. Histochem. Cytochem. 38, 573–580 (1990).

    Article  CAS  PubMed  Google Scholar 

  9. Kovacs, A. L., Palfia, Z., Rez, G., Vellai, T. & Kovacs, J. Sequestration revisited: integrating traditional electron microscopy, de novo assembly and new results. Autophagy 3, 655–662 (2007).

    Article  PubMed  Google Scholar 

  10. Axe, E. L. et al. Autophagosome formation from membrane compartments enriched in phosphatidylinositol 3-phosphate and dynamically connected to the endoplasmic reticulum. J. Cell Biol. 182, 685–701 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Walker, S., Chandra, P., Manifava, M., Axe, E. & Ktistakis, N. T. Making autophagosomes: localized synthesis of phosphatidylinositol 3-phosphate holds the clue. Autophagy 4, 1093–1096 (2008).

    Article  CAS  PubMed  Google Scholar 

  12. Fujita, N. et al. An Atg4B mutant hampers the lipidation of LC3 paralogues and causes defects in autophagosome closure. Mol. Biol. Cell 19, 4651–4659 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Fengsrud, M. et al. Ultrastructural and immunocytochemical characterization of autophagic vacuoles in isolated hepatocytes: effects of vinblastine and asparagine on vacuole distributions. Exp. Cell Res. 221, 504–519 (1995).

    Article  CAS  PubMed  Google Scholar 

  14. Mizushima, N. et al. Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J. Cell Biol. 152, 657–668 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Mizushima, N. et al. Mouse Apg16L, a novel WD-repeat protein, targets to the autophagic isolation membrane with the Apg12–Apg5 conjugate. J. Cell Sci. 116, 1679–1688 (2003).

    Article  CAS  PubMed  Google Scholar 

  16. Saitoh, T. et al. Loss of the autophagy protein Atg16L1 enhances endotoxin-induced IL-1β production. Nature 456, 264–268 (2008).

    Article  CAS  PubMed  Google Scholar 

  17. McIntosh, R., Nicastro, D. & Mastronarde, D. New views of cells in 3D: an introduction to electron tomography. Trends Cell Biol. 15, 43–51 (2005).

    Article  CAS  PubMed  Google Scholar 

  18. Bannykh, S. I., Rowe, T. & Balch, W. E. The organization of endoplasmic reticulum export complexes. J. Cell Biol. 135, 19–35 (1996).

    Article  CAS  PubMed  Google Scholar 

  19. Saitoh, T. et al. TWEAK induces NF-kappaB2 p100 processing and long lasting NF-kappaB activation. J. Biol. Chem. 278, 36005–36012 (2003).

    Article  CAS  PubMed  Google Scholar 

  20. Mastronarde, D. N. Dual-axis tomography: an approach with alignment methods that preserve resolution. J. Struct. Biol. 120, 343–352 (1997).

    Article  CAS  PubMed  Google Scholar 

  21. Kremer, J. R., Mastronarde, D. N. & McIntosh, J. R. Computer visualization of three-dimensional image data using IMOD. J. Struct. Biol. 116, 71–76 (1996).

    Article  CAS  PubMed  Google Scholar 

  22. Wakana, Y. et al. Bap31 is an itinerant protein that moves between the peripheral endoplasmic reticulum (ER) and a juxtanuclear compartment related to ER-associated Degradation. Mol. Biol. Cell 19, 1825–1836 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references


We thank H. Nishioka and N. Kajimura for technical assistance with electron tomography, K. Nishino for assistance with setting up a computer and for useful comments on figure presentations, H. Omori for technical assistance with immunoelectron microscopy and K. Matsunaga and N. Taguchi for technical assistance with plasmid construction. This study was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

Author information

Authors and Affiliations



M.H.N., N.F., T.N., T.Y. and A.Y. designed experiments. M.H.N. performed conventional electron microscopy and electron tomography, N.F. performed cell biological and immuofluorescence experiments and A.Y. performed immunoelectron microscopy. M.H.N., T.Y. and A.Y. wrote the manuscript. T.N. and A.Y. contributed to conceptual discussions. A.Y. and T.Y. supervised the project.

Corresponding authors

Correspondence to Tamotsu Yoshimori or Akitsugu Yamamoto.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 3010 kb)

Supplementary Movie 1

Supplementary Movie 1 (MOV 5714 kb)

Supplementary Movie 2

Supplementary Movie 2 (MOV 1100 kb)

Supplementary Movie 3

Supplementary Movie 3 (MOV 2845 kb)

Supplementary Movie 4

Supplementary Movie 4 (MOV 2014 kb)

Supplementary Movie 5

Supplementary Movie 5 (MOV 1487 kb)

Supplementary Movie 6

Supplementary Movie 6 (MOV 1758 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Hayashi-Nishino, M., Fujita, N., Noda, T. et al. A subdomain of the endoplasmic reticulum forms a cradle for autophagosome formation. Nat Cell Biol 11, 1433–1437 (2009).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


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