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Individual components of paired typical NLR immune receptors are regulated by distinct E3 ligases

Nature Plants volume 4, pages 699–710 (2018)Cite this article

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Abstract

In plants and animals, nucleotide-binding leucine-rich repeat (NLR) proteins serve as intracellular immune receptors. Defence signalling by NLRs often requires the formation of NLR heteropairs. Our knowledge of the molecular mechanism regulating this process is limited. In a reverse genetic screen to identify the partner of the Arabidopsis typical NLR, SUPRESSOR OF NPR1, CONSTITUTIVE 1 (SNC1), we discovered three NLRs that are redundantly required for SNC1-mediated defence, which were named SIDEKICK SNC1 1 (SIKIC1), SIKIC2 and SIKIC3. Immunoprecipitation–mass spectrometry analyses revealed that SIKIC2 physically associates with SNC1. We also uncovered that the protein level of SIKIC2 is under the control of two previously uncharacterized redundant E3 ubiquitin ligases MUSE1 and MUSE2. As SNC1 accumulation has previously been shown to be regulated by the E3 ubiquitin ligase SCFCPR1, this report provides evidence that the homeostasis of individual components of partnered typical NLRs is subjected to differential regulation via ubiquitin-mediated protein degradation.

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Fig. 1: SIKIC1, SIKIC2 and SIKIC3 are redundantly required for snc1-mediated autoimmunity.
Fig. 2: muse1-1 enhances immunity in the mos4 snc1 and snc1 background.
Fig. 3: MUSE1 exhibits E3 ubiquitin ligase activity.
Fig. 4: MUSE1 and MUSE2 have overlapping functions.
Fig. 5: MUSE1 and MUSE2 are specifically involved in SNC1-mediated immunity.
Fig. 6: MUSE1 and MUSE2 negatively regulate the accumulation of SIKIC proteins.

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Acknowledgements

We thank Q. Xie (Institute of Genetics and Developmental Biology, Chinese Academy of Sciences) for sharing the pET28-AtUBA2-His construct, X. Deng (Peking University/Yale University) for sharing the pGEX-GST-AtUBC8 construct for protein expression, J. Stuttmann (Martin Luther University, Halle-Wittenberg) for sharing the seeds of N. benthaniana eds1 and K. Kirchsteiger and M. Estelle (UCSD) for offering insights in the in vitro ubiquitination assay. Y. Li (Institute of Genetics and Developmental Biology, Chinese Academy of Sciences) is thanked for Illumina sequencing data analyses. Y. Qiu (University of British Columbia) is thanked for helping with the phylogenetic analyses of MUSE1. We are grateful to N. Zheng and W. Xing (University of Washington) for enlightening discussions about MUSE1 secondary structure prediction and E3 functions. The Arabidopsis Biological Resource Center is acknowledged for providing the muse1-7 and muse2-1 knockout T-DNA mutant seeds. This work was financially supported by the Natural Sciences and Engineering Research Council (NSERC) Discovery program of Canada and Canadian Foundation for Innovation (CFI) (to X.L.), the Chinese Scholarship Council (to Y.W., Y.H., S.X. and X.C.), the Dewar Memorial fund (to X.L.), NSERC Graduate scholarships (to K.A. and K.C.M.J.) and the UBC Four-Year Fellowship (to F.X., K.C.M.J. and O.X.D.).

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Author notes

  1. Oliver Xiaoou Dong

    Present address: Department of Plant Pathology, University of California, Davis, Davis, CA, USA

  2. Fang Xu

    Present address: Cold Spring Harbor Laboratory, Cold Spring Harbor, New York, NY, USA

  3. These authors contributed equally to this work: Oliver Xiaoou Dong, Kevin Ao, Fang Xu.

Authors and Affiliations

  1. Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, Canada

    Oliver Xiaoou Dong, Kevin Ao, Fang Xu, Kaeli C. M. Johnson, Yuxiang Wu, Shitou Xia, Yanan Liu, Yan Huang, Xuejin Chen & Xin Li

  2. Department of Botany, University of British Columbia, Vancouver, British Columbia, Canada

    Oliver Xiaoou Dong, Kevin Ao, Fang Xu, Kaeli C. M. Johnson, Yanan Liu, Yan Huang, Yuelin Zhang & Xin Li

  3. College of Agriculture, Shanxi Agriculture University, Jinzhong, China

    Yuxiang Wu

  4. National Institute of Biological Sciences, Beijing, China

    Lin Li

  5. Hunan Provincial Key Laboratory of Phytohormones and Growth Development, Hunan Agricultural University, Changsha, China

    Shitou Xia & She Chen

  6. College of Life Science, Sichuan Agricultural University, Ya’an, China

    Yan Huang

  7. Department of Biology, University of Copenhagen, Copenhagen, Denmark

    Eleazar Rodriguez & Morten Petersen

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Contributions

O.X.D. and K.A. performed most of the described experiments under the supervision of X.L. F.X. performed the SNC1/snc1 IP. Y.W. performed the crude mapping of muse1-1. L.L. performed the MS analyses. S.C. and Y.Z. analysed the MS and Illumina data sets. K.C.M.J., Y.H., S.X. and X.C. mapped and identified five additional mutant alleles of muse1. Y.L. constructed the library for Illumina sequencing. E.R. and M.P. shared their dominant-negative TNL collection. O.X.D. and X.L. wrote the manuscript. All authors reviewed the manuscript.

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Correspondence to Xin Li.

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Dong, O.X., Ao, K., Xu, F. et al. Individual components of paired typical NLR immune receptors are regulated by distinct E3 ligases. Nature Plants 4, 699–710 (2018). https://doi.org/10.1038/s41477-018-0216-8

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