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The Arabidopsis O-fucosyltransferase SPINDLY activates nuclear growth repressor DELLA

Nature Chemical Biology volume 13, pages 479–485 (2017)Cite this article

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Abstract

Plant development requires coordination among complex signaling networks to enhance the plant's adaptation to changing environments. DELLAs, transcription regulators originally identified as repressors of phytohormone gibberellin signaling, play a central role in integrating multiple signaling activities via direct protein interactions with key transcription factors. Here, we found that DELLA is mono-O-fucosylated by the novel O-fucosyltransferase SPINDLY (SPY) in Arabidopsis thaliana. O-fucosylation activates DELLA by promoting its interaction with key regulators in brassinosteroid- and light-signaling pathways, including BRASSINAZOLE-RESISTANT1 (BZR1), PHYTOCHROME-INTERACTING-FACTOR3 (PIF3) and PIF4. Moreover, spy mutants displayed elevated responses to gibberellin and brassinosteroid, and increased expression of common target genes of DELLAs, BZR1 and PIFs. Our study revealed that SPY-dependent protein O-fucosylation plays a key role in regulating plant development. This finding may have broader importance because SPY orthologs are conserved in prokaryotes and eukaryotes, thus suggesting that intracellular O-fucosylation may regulate a wide range of biological processes in diverse organisms.

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Figure 1: RGA shows SPY-dependent O-fucosylation in planta.
Figure 2: SPY is a novel protein O-fucosyltransferase (POFUT).
Figure 3: Phenotype and enzyme-activity analyses of the spy mutants.
Figure 4: O-fucosylation enhances RGA activity by promoting RGA binding to its interactors.
Figure 5: SPY and SEC compete with each other in reciprocal modifications of RGA.
Figure 6: Model of the role of O-fucosylation versus O-GlcNAcylation of RGA.

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References

  1. Vert, G. & Chory, J. Crosstalk in cellular signaling: background noise or the real thing? Dev. Cell 21, 985–991 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Silverstone, A.L., Ciampaglio, C.N. & Sun, T. The Arabidopsis RGA gene encodes a transcriptional regulator repressing the gibberellin signal transduction pathway. Plant Cell 10, 155–169 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Peng, J. et al. The Arabidopsis GAI gene defines a signaling pathway that negatively regulates gibberellin responses. Genes Dev. 11, 3194–3205 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Sun, T.P. The molecular mechanism and evolution of the GA-GID1-DELLA signaling module in plants. Curr. Biol. 21, R338–R345 (2011).

    Article  CAS  PubMed  Google Scholar 

  5. Davière, J.M. & Achard, P. Gibberellin signaling in plants. Development 140, 1147–1151 (2013).

    Article  PubMed  CAS  Google Scholar 

  6. de Lucas, M. et al. A molecular framework for light and gibberellin control of cell elongation. Nature 451, 480–484 (2008).

    Article  CAS  PubMed  Google Scholar 

  7. Feng, S. et al. Coordinated regulation of Arabidopsis thaliana development by light and gibberellins. Nature 451, 475–479 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Xu, H., Liu, Q., Yao, T. & Fu, X. Shedding light on integrative GA signaling. Curr. Opin. Plant Biol. 21, 89–95 (2014).

    Article  CAS  PubMed  Google Scholar 

  9. Ueguchi-Tanaka, M. et al. GIBBERELLIN INSENSITIVE DWARF1 encodes a soluble receptor for gibberellin. Nature 437, 693–698 (2005).

    Article  CAS  PubMed  Google Scholar 

  10. Murase, K., Hirano, Y., Sun, T.P. & Hakoshima, T. Gibberellin-induced DELLA recognition by the gibberellin receptor GID1. Nature 456, 459–463 (2008).

    Article  CAS  PubMed  Google Scholar 

  11. McGinnis, K.M. et al. The Arabidopsis SLEEPY1 gene encodes a putative F-box subunit of an SCF E3 ubiquitin ligase. Plant Cell 15, 1120–1130 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Sasaki, A. et al. Accumulation of phosphorylated repressor for gibberellin signaling in an F-box mutant. Science 299, 1896–1898 (2003).

    Article  CAS  PubMed  Google Scholar 

  13. Griffiths, J. et al. Genetic characterization and functional analysis of the GID1 gibberellin receptors in Arabidopsis. Plant Cell 18, 3399–3414 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Dill, A., Jung, H.-S. & Sun, T.P. The DELLA motif is essential for gibberellin-induced degradation of RGA. Proc. Natl. Acad. Sci. USA 98, 14162–14167 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Silverstone, A.L. et al. Functional analysis of SPINDLY in gibberellin signaling in Arabidopsis. Plant Physiol. 143, 987–1000 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Wilson, R.N. & Somerville, C.R. Phenotypic suppression of the gibberellin-insensitive mutant (gai) of Arabidopsis. Plant Physiol. 108, 495–502 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Olszewski, N.E., West, C.M., Sassi, S.O. & Hartweck, L.M. O-GlcNAc protein modification in plants: evolution and function. Biochim. Biophys. Acta 1800, 49–56 (2010).

    Article  CAS  PubMed  Google Scholar 

  18. Hartweck, L.M., Scott, C.L. & Olszewski, N.E. Two O-linked N-acetylglucosamine transferase genes of Arabidopsis thaliana L. Heynh. have overlapping functions necessary for gamete and seed development. Genetics 161, 1279–1291 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Jacobsen, S.E., Binkowski, K.A. & Olszewski, N.E. SPINDLY, a tetratricopeptide repeat protein involved in gibberellin signal transduction in Arabidopsis. Proc. Natl. Acad. Sci. USA 93, 9292–9296 (1996).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Zentella, R. et al. O-GlcNAcylation of master growth repressor DELLA by SECRET AGENT modulates multiple signaling pathways in Arabidopsis. Genes Dev. 30, 164–176 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Jacobsen, S.E. & Olszewski, N.E. Mutations at the SPINDLY locus of Arabidopsis alter gibberellin signal transduction. Plant Cell 5, 887–896 (1993).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Luther, K.B. & Haltiwanger, R.S. Role of unusual O-glycans in intercellular signaling. Int. J. Biochem. Cell Biol. 41, 1011–1024 (2009).

    Article  CAS  PubMed  Google Scholar 

  23. Okajima, T. & Irvine, K.D. Regulation of notch signaling by O-linked fucose. Cell 111, 893–904 (2002).

    Article  CAS  PubMed  Google Scholar 

  24. Hofsteenge, J. et al. C-mannosylation and O-fucosylation of the thrombospondin type 1 module. J. Biol. Chem. 276, 6485–6498 (2001).

    Article  CAS  PubMed  Google Scholar 

  25. Lira-Navarrete, E. et al. Structural insights into the mechanism of protein O-fucosylation. PLoS One 6, e25365 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Chen, C.I. et al. Structure of human POFUT2: insights into thrombospondin type 1 repeat fold and O-fucosylation. EMBO J. 31, 3183–3197 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Lazarus, M.B., Nam, Y., Jiang, J., Sliz, P. & Walker, S. Structure of human O-GlcNAc transferase and its complex with a peptide substrate. Nature 469, 564–567 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Kreppel, L.K. & Hart, G.W. Regulation of a cytosolic and nuclear O-GlcNAc transferase: role of the tetratricopeptide repeats. J. Biol. Chem. 274, 32015–32022 (1999).

    Article  CAS  PubMed  Google Scholar 

  29. Wu, Z.L., Ethen, C.M., Prather, B., Machacek, M. & Jiang, W. Universal phosphatase-coupled glycosyltransferase assay. Glycobiology 21, 727–733 (2011).

    Article  CAS  PubMed  Google Scholar 

  30. Baykov, A.A., Evtushenko, O.A. & Avaeva, S.M. A malachite green procedure for orthophosphate determination and its use in alkaline phosphatase-based enzyme immunoassay. Anal. Biochem. 171, 266–270 (1988).

    Article  CAS  PubMed  Google Scholar 

  31. Lazarus, M.B. et al. HCF-1 is cleaved in the active site of O-GlcNAc transferase. Science 342, 1235–1239 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Schimpl, M. et al. O-GlcNAc transferase invokes nucleotide sugar pyrophosphate participation in catalysis. Nat. Chem. Biol. 8, 969–974 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Martinez-Fleites, C. et al. Structure of an O-GlcNAc transferase homolog provides insight into intracellular glycosylation. Nat. Struct. Mol. Biol. 15, 764–765 (2008).

    Article  CAS  PubMed  Google Scholar 

  34. Bai, M.Y. et al. Brassinosteroid, gibberellin, and phytochrome impinge on a common transcription module in Arabidopsis. Nat. Cell Biol. 14, 810–817 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Oh, E. et al. Cell elongation is regulated through a central circuit of interacting transcription factors in the Arabidopsis hypocotyl. eLife 3, e03031 (2014).

    Article  PubMed Central  CAS  Google Scholar 

  36. Gallego-Bartolomé, J. et al. Molecular mechanism for the interaction between gibberellin and brassinosteroid signaling pathways in Arabidopsis. Proc. Natl. Acad. Sci. USA 109, 13446–13451 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  37. Nozue, K. et al. Rhythmic growth explained by coincidence between internal and external cues. Nature 448, 358–361 (2007).

    Article  CAS  PubMed  Google Scholar 

  38. Soy, J. et al. Phytochrome-imposed oscillations in PIF3 protein abundance regulate hypocotyl growth under diurnal light/dark conditions in Arabidopsis. Plant J. 71, 390–401 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Silverstone, A.L., Mak, P.Y.A., Martínez, E.C. & Sun, T.P. The new RGA locus encodes a negative regulator of gibberellin response in Arabidopsis thaliana. Genetics 146, 1087–1099 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Hart, G.W., Slawson, C., Ramirez-Correa, G. & Lagerlof, O. Cross talk between O-GlcNAcylation and phosphorylation: roles in signaling, transcription, and chronic disease. Annu. Rev. Biochem. 80, 825–858 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Bond, M.R. & Hanover, J.A. A little sugar goes a long way: the cell biology of O-GlcNAc. J. Cell Biol. 208, 869–880 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Greenboim-Wainberg, Y. et al. Cross talk between gibberellin and cytokinin: the Arabidopsis GA response inhibitor SPINDLY plays a positive role in cytokinin signaling. Plant Cell 17, 92–102 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Tseng, T.S., Salomé, P.A., McClung, C.R. & Olszewski, N.E. SPINDLY and GIGANTEA interact and act in Arabidopsis thaliana pathways involved in light responses, flowering, and rhythms in cotyledon movements. Plant Cell 16, 1550–1563 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Zentella, R. et al. Global analysis of DELLA direct targets in early gibberellin signaling in Arabidopsis. Plant Cell 19, 3037–3057 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Earley, K.W. et al. Gateway-compatible vectors for plant functional genomics and proteomics. Plant J. 45, 616–629 (2006).

    Article  CAS  PubMed  Google Scholar 

  46. Dill, A., Thomas, S.G., Hu, J., Steber, C.M. & Sun, T.P. The Arabidopsis F-box protein SLEEPY1 targets gibberellin signaling repressors for gibberellin-induced degradation. Plant Cell 16, 1392–1405 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Zhang, Z.L. et al. Scarecrow-like 3 promotes gibberellin signaling by antagonizing master growth repressor DELLA in Arabidopsis. Proc. Natl. Acad. Sci. USA 108, 2160–2165 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Silverstone, A.L. et al. Repressing a repressor: gibberellin-induced rapid reduction of the RGA protein in Arabidopsis. Plant Cell 13, 1555–1566 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Swain, S.M., Tseng, T.S., Thornton, T.M., Gopalraj, M. & Olszewski, N.E. SPINDLY is a nuclear-localized repressor of gibberellin signal transduction expressed throughout the plant. Plant Physiol. 129, 605–615 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Udeshi, N.D., Compton, P.D., Shabanowitz, J., Hunt, D.F. & Rose, K.L. Methods for analyzing peptides and proteins on a chromatographic timescale by electron-transfer dissociation mass spectrometry. Nat. Protoc. 3, 1709–1717 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Berk, J.M. et al. O-linked β-N-acetylglucosamine (O-GlcNAc) regulates emerin binding to barrier to autointegration factor (BAF) in a chromatin- and lamin B-enriched “niche”. J. Biol. Chem. 288, 30192–30209 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Xu, Y., Strickland, E.C. & Fitzgerald, M.C. Thermodynamic analysis of protein folding and stability using a tryptophan modification protocol. Anal. Chem. 86, 7041–7048 (2014).

    Article  CAS  PubMed  Google Scholar 

  53. Arnold, K., Bordoli, L., Kopp, J. & Schwede, T. The SWISS-MODEL workspace: a web-based environment for protein structure homology modelling. Bioinformatics 22, 195–201 (2006).

    Article  CAS  PubMed  Google Scholar 

  54. Biasini, M. et al. SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Res. 42, W252–W258 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank C. Toleman for assistance with in vitro enzyme assays and G. Dubay at Duke Chemistry Instrument Facilities for help with the MALDI–MS analysis. We also thank Z.-M. Pei and J. Siedow for helpful comments on the manuscript. This work was supported by the National Institutes of Health (R01 GM100051 to T.-p.S.; R01 GM037537 to D.F.H.), the National Science Foundation (MCB-0923723 to T.-p.S.; MCB-0516690, MCB-0820666 and MCB-1158089 to N.E.O.), the Department of Energy (DE-SC0014077 to T.-p.S.) and the US Department of Agriculture (2014-67013-21548 to T.-p.S.).

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Author notes

  1. Rodolfo Zentella and Ning Sui: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biology, Duke University, Durham, North Carolina, USA

    Rodolfo Zentella, Ning Sui, Wen-Ping Hsieh, Jianhong Hu & Tai-ping Sun

  2. Department of Chemistry, University of Virginia, Charlottesville, Virginia, USA

    Benjamin Barnhill, Jeffrey Shabanowitz & Donald F Hunt

  3. Department of Biochemistry, Duke University Medical Center, Durham, North Carolina, USA

    Michael Boyce & Pei Zhou

  4. Department of Plant Biology, University of Minnesota, Saint Paul, Minnesota, USA

    Neil E Olszewski

  5. Department of Chemistry, Duke University, Durham, North Carolina, USA

    Pei Zhou

  6. Department of Pathology, University of Virginia, Charlottesville, Virginia, USA

    Donald F Hunt

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Contributions

R.Z., N.S. and T.-p.S. conceived and designed the research project. R.Z., N.S., W.-P.H. and J.H. performed molecular biology and biochemical analysis; R.Z., N.S., P.Z. and T.-p.S. analyzed data. B.B. performed LC–ETD–MS/MS analysis, and B.B., J.S. and D.F.H. analyzed MS data. P.Z. also provided advice on protein purification and structure modeling. M.B. provided reagents and advice for in vitro enzyme assays with MALDI–MS; N.E.O. provided experimental materials and shared unpublished results. R.Z., N.S. and T.-p.S. wrote the manuscript.

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Correspondence to Tai-ping Sun.

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Zentella, R., Sui, N., Barnhill, B. et al. The Arabidopsis O-fucosyltransferase SPINDLY activates nuclear growth repressor DELLA. Nat Chem Biol 13, 479–485 (2017). https://doi.org/10.1038/nchembio.2320

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