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Noncanonical recognition and UBL loading of distinct E2s by autophagy-essential Atg7

Nature Structural & Molecular Biology volume 19, pages 1250–1256 (2012)Cite this article

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Abstract

Autophagy requires ubiquitin-like Atg8 and Atg12 conjugation systems, where Atg7 has a critical role as the sole E1 enzyme. Although Atg7 recognizes two distinct E2s, Atg3 and Atg10, it is not understood how Atg7 correctly loads these E2s with their cognate ubiquitin-like proteins, Atg8 and Atg12. Here, we report the crystal structures of the N-terminal domain of Atg7 bound to Atg10 or Atg3 of thermotolerant yeast and plant homologs. The observed Atg7-Atg10 and Atg7-Atg3 interactions, which resemble each other but are quite distinct from the canonical E1-E2 interaction, makes Atg7 suitable for transferring Atg12 to Atg10 and Atg8 to Atg3 by a trans mechanism. Notably, in vitro experiments showed that Atg7 loads Atg3 and Atg10 with Atg8 and Atg12 in a nonspecific manner, which suggests that cognate conjugate formation in vivo is not an intrinsic quality of Atg7.

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Figure 1: Structure of the KmAtg7NTD–KmAtg10 complex and identification of the important residues for the transthioesterification reaction.
Figure 2: Structure of the AtAtg7NTD–AtAtg3 complex and identification of the important residues for the transthioesterification reaction.
Figure 3: Atg12 transfer from Atg7 to Atg10 proceeds in trans.
Figure 4: Nonselective UBL loading of Atg3 and Atg10 by Atg7.

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Acknowledgements

The synchrotron radiation experiments were performed at beamlines BL41XU of SPring8 and BL5A, NE3A and NW12A of KEK in Japan. This work was supported in part by JSPS KAKENHI grant numbers 23687012 (to N.N.N.), 23000015 (to Y.O.), MEXT KAKENHI 24113725 (to N.N.N.) and MEXT Targeted Proteins Research Program (to F.I. and Y.O.).

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Author notes

  1. Masaya Yamaguchi and Kazuaki Matoba: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Structural Biology, Faculty of Advanced Life Science, Hokkaido University, Sapporo, Japan

    Masaya Yamaguchi, Ryoko Sawada, Yoshihiro Kobashigawa & Fuyuhiko Inagaki

  2. Institute of Microbial Chemistry, Tokyo, Japan

    Kazuaki Matoba, Yuko Fujioka & Nobuo N Noda

  3. Integrated Research Institute, Tokyo Institute of Technology, Yokohama, Japan

    Hitoshi Nakatogawa, Hayashi Yamamoto & Yoshinori Ohsumi

  4. Department of Applied Molecular Bioscience, Yamaguchi University Graduate School of Medicine, Ube, Japan

    Hisashi Hoshida & Rinji Akada

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  1. Masaya Yamaguchi
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Contributions

M.Y., K.M. and N.N.N. performed structural studies; M.Y., K.M., R.S., Y.F. and Y.K. performed biochemical studies; H.N. and H.Y. performed yeast experiments; H.H. and R.A. cloned Atg homologs from K. marxianus. M.Y., K.M., Y.O., N.N.N. and F.I. analyzed data and wrote the manuscript. All authors discussed the results and commented on the manuscript. N.N.N. and F.I. supervised the work.

Corresponding authors

Correspondence to Nobuo N Noda or Fuyuhiko Inagaki.

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Yamaguchi, M., Matoba, K., Sawada, R. et al. Noncanonical recognition and UBL loading of distinct E2s by autophagy-essential Atg7. Nat Struct Mol Biol 19, 1250–1256 (2012). https://doi.org/10.1038/nsmb.2451

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