Two distinct mechanisms target the autophagy-related E3 complex to the pre-autophagosomal structure

Abstract

In autophagy, Atg proteins organize the pre-autophagosomal structure (PAS) to initiate autophagosome formation. Previous studies in yeast revealed that the autophagy-related E3 complex Atg12-Atg5-Atg16 is recruited to the PAS via Atg16 interaction with Atg21, which binds phosphatidylinositol 3-phosphate (PI3P) produced at the PAS, to stimulate conjugation of the ubiquitin-like protein Atg8 to phosphatidylethanolamine. Here, we discover a novel mechanism for the PAS targeting of Atg12-Atg5-Atg16, which is mediated by the interaction of Atg12 with the Atg1 kinase complex that serves as a scaffold for PAS organization. While autophagy is partially defective without one of these mechanisms, cells lacking both completely lose the PAS localization of Atg12-Atg5-Atg16 and show no autophagic activity. As with the PI3P-dependent mechanism, Atg12-Atg5-Atg16 recruited via the Atg12-dependent mechanism stimulates Atg8 lipidation, but also has the specific function of facilitating PAS scaffold assembly. Thus, this study significantly advances our understanding of the nucleation step in autophagosome formation.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files.

Article and author information

Author details

  1. Kumi Harada

    School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  2. Tetsuya Kotani

    School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  3. Hiromi Kirisako

    School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  4. Machiko Sakoh-Nakatogawa

    School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  5. Yu Oikawa

    Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  6. Yayoi Kimura

    Advanced Medical Research Center, Yokohama City University, Yokohama, Japan
    Competing interests
    No competing interests declared.
  7. Hisashi Hirano

    Advanced Medical Research Center, Yokohama City University, Yokohama, Japan
    Competing interests
    No competing interests declared.
  8. Hayashi Yamamoto

    Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  9. Yoshinori Ohsumi

    Institute of Innovative Research, Tokyo Institute of Technology, Yokohama, Japan
    Competing interests
    No competing interests declared.
  10. Hitoshi Nakatogawa

    School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
    For correspondence
    hnakatogawa@bio.titech.ac.jp
    Competing interests
    Hitoshi Nakatogawa, Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5828-0741

Funding

Ministry of Education, Culture, Sports, Science, and Technology (25111003)

  • Hitoshi Nakatogawa

Ministry of Education, Culture, Sports, Science, and Technology (17H01430)

  • Hitoshi Nakatogawa

Ministry of Education, Culture, Sports, Science, and Technology (23000015)

  • Yoshinori Ohsumi

Japan Science and Technology Agency (JPMJCR13M7)

  • Hitoshi Nakatogawa

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2019, Harada et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 4,400
    views
  • 725
    downloads
  • 57
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Kumi Harada
  2. Tetsuya Kotani
  3. Hiromi Kirisako
  4. Machiko Sakoh-Nakatogawa
  5. Yu Oikawa
  6. Yayoi Kimura
  7. Hisashi Hirano
  8. Hayashi Yamamoto
  9. Yoshinori Ohsumi
  10. Hitoshi Nakatogawa
(2019)
Two distinct mechanisms target the autophagy-related E3 complex to the pre-autophagosomal structure
eLife 8:e43088.
https://doi.org/10.7554/eLife.43088

Share this article

https://doi.org/10.7554/eLife.43088

Further reading

    1. Cell Biology
    2. Neuroscience
    Luis Sánchez-Guardado, Peyman Callejas Razavi ... Carlos Lois
    Research Article

    The assembly and maintenance of neural circuits is crucial for proper brain function. Although the assembly of brain circuits has been extensively studied, much less is understood about the mechanisms controlling their maintenance as animals mature. In the olfactory system, the axons of olfactory sensory neurons (OSNs) expressing the same odor receptor converge into discrete synaptic structures of the olfactory bulb (OB) called glomeruli, forming a stereotypic odor map. The OB projection neurons, called mitral and tufted cells (M/Ts), have a single dendrite that branches into a single glomerulus, where they make synapses with OSNs. We used a genetic method to progressively eliminate the vast majority of M/T cells in early postnatal mice, and observed that the assembly of the OB bulb circuits proceeded normally. However, as the animals became adults the apical dendrite of remaining M/Ts grew multiple branches that innervated several glomeruli, and OSNs expressing single odor receptors projected their axons into multiple glomeruli, disrupting the olfactory sensory map. Moreover, ablating the M/Ts in adult animals also resulted in similar structural changes in the projections of remaining M/Ts and axons from OSNs. Interestingly, the ability of these mice to detect odors was relatively preserved despite only having 1–5% of projection neurons transmitting odorant information to the brain, and having highly disrupted circuits in the OB. These results indicate that a reduced number of projection neurons does not affect the normal assembly of the olfactory circuit, but induces structural instability of the olfactory circuitry of adult animals.

    1. Cell Biology
    2. Developmental Biology
    Yi Sun, Zhe Chen ... Chengtian Zhao
    Short Report

    How cells regulate the size of their organelles remains a fundamental question in cell biology. Cilia, with their simple structure and surface localization, provide an ideal model for investigating organelle size control. However, most studies on cilia length regulation are primarily performed on several single-celled organisms. In contrast, the mechanism of length regulation in cilia across diverse cell types within multicellular organisms remains a mystery. Similar to humans, zebrafish contain diverse types of cilia with variable lengths. Taking advantage of the transparency of zebrafish embryos, we conducted a comprehensive investigation into intraflagellar transport (IFT), an essential process for ciliogenesis. By generating a transgenic line carrying Ift88-GFP transgene, we observed IFT in multiple types of cilia with varying lengths. Remarkably, cilia exhibited variable IFT speeds in different cell types, with longer cilia exhibiting faster IFT speeds. This increased IFT speed in longer cilia is likely not due to changes in common factors that regulate IFT, such as motor selection, BBSome proteins, or tubulin modification. Interestingly, longer cilia in the ear cristae tend to form larger IFT compared to shorter spinal cord cilia. Reducing the size of IFT particles by knocking down Ift88 slowed IFT speed and resulted in the formation of shorter cilia. Our study proposes an intriguing model of cilia length regulation via controlling IFT speed through the modulation of the size of the IFT complex. This discovery may provide further insights into our understanding of how organelle size is regulated in higher vertebrates.