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The crystal structure of Deg9 reveals a novel octameric-type HtrA protease

Nature Plants volume 3, pages 973–982 (2017)Cite this article

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Abstract

The high temperature requirement A (HtrA) proteases (also termed Deg proteases) play important roles in diverse organisms by regulating protein quality and quantity. One of the 16 Arabidopsis homologs, Deg9, is located in the nucleus where it modulates cytokinin- and light-mediated signalling via degrading the ARABIDOPSIS RESPONSE REGULATOR 4 (ARR4). To uncover the structural features underlying the proteolytic activity of Deg9, we determined its crystal structure. Unlike the well-established trimeric building block of HtrAs, Deg9 displays a novel octameric structure consisting of two tetrameric rings that have distinct conformations. Based on the structural architecture, we generated several mutant variants of Deg9, determined their structure and tested their proteolytic activity towards ARR4. The results of the structural and biochemical analyses allowed us to propose a model for a novel mechanism of substrate recognition and activity regulation of Deg9. In this model, protease activation of one tetramer is mediated by en-bloc reorientation of the protease domains to open an entrance for the substrate in the opposite (inactive) tetramer. This study provides the structural basis for understanding how the levels of nuclear signal components are regulated by a plant protease.

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Fig. 1: Structure of the Deg9 octamer.
Fig. 2: Structures of the Deg9 variant lacking the PDZ domain and a Deg9 substitute fusion protein.
Fig. 3: Conformational differences in the activation domains between tetramers A and B.
Fig. 4: Heterogeneous dynamics in the octamer at 4 °C and 16 °C.
Fig. 5: Intersubunit crosstalk between tetramers.
Fig. 6: The model of Deg9 protease activation and substrate access.

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Acknowledgements

This study was supported by the Joint NSFC-ISF Research Program (jointly funded by the National Natural Science Foundation of China and the Israel Science Foundation (31661143026)), the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB17000000) and the Youth Innovation Promotion Association CAS (2013059). We thank the Shanghai Synchrotron Radiation Facility and the Lin Liu lab for technical support during data collection and analysis.

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  1. Min Ouyang and Xiaoyi Li contributed equally to this work.

Authors and Affiliations

  1. Photosynthesis Research Center, Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Beijing, China

    Min Ouyang, Xiaoyi Li, Shun Zhao, Hua Pu, Jianren Shen & Lixin Zhang

  2. Research Institute for Interdisciplinary Science and Graduate School of Natural Science and Technology, Okayama University, Okayama, Japan

    Jianren Shen

  3. The Robert H. Smith Institute of Plant Sciences and Genetics in Agriculture, The Hebrew University of Jerusalem, Rehovot, Israel

    Zach Adam

  4. Research Institute of Molecular Pathology (IMP), Vienna, Austria

    Tim Clausen

  5. College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China

    Lixin Zhang

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Contributions

M.O. and L.Z. designed the study; M.O. and X.L. performed the research; M.O., X.L., T.C., S.Z. and H.P. analysed the data; M.O. and L.Z. wrote the paper. J.S., T.C. and Z.A. assisted in writing and discussion.

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Correspondence to Lixin Zhang.

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Ouyang, M., Li, X., Zhao, S. et al. The crystal structure of Deg9 reveals a novel octameric-type HtrA protease. Nature Plants 3, 973–982 (2017). https://doi.org/10.1038/s41477-017-0060-2

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