A secreted LysM effector protects fungal hyphae through chitin-dependent homodimer polymerization

Autoři: Andrea Sánchez-Vallet aff001;  Hui Tian aff001;  Luis Rodriguez-Moreno aff001;  Dirk-Jan Valkenburg aff001;  Raspudin Saleem-Batcha aff003;  Stephan Wawra aff004;  Anja Kombrink aff001;  Leonie Verhage aff001;  Ronnie de Jonge aff001;  H. Peter van Esse aff001;  Alga Zuccaro aff004;  Daniel Croll aff005;  Jeroen R. Mesters aff003;  Bart P. H. J. Thomma aff001
Působiště autorů: Laboratory of Phytopathology, Wageningen University& Research, Wageningen, The Netherlands aff001;  Plant Pathology Group, Institute of Integrative Biology, ETH Zurich, Zurich, Switzerland aff002;  Institute of Biochemistry, Center for Structural and Cell Biology in Medicine, University of Lübeck, Lübeck, Germany aff003;  University of Cologne, Institute for Plant Sciences, Cluster of Excellence on Plant Sciences (CEPLAS), Cologne, Germany aff004;  Institute of Biology, University of Neuchâtel, Neuchâtel, Switzerland aff005
Vyšlo v časopise: A secreted LysM effector protects fungal hyphae through chitin-dependent homodimer polymerization. PLoS Pathog 16(6): e32767. doi:10.1371/journal.ppat.1008652
Kategorie: Research Article
doi: 10.1371/journal.ppat.1008652


Plants trigger immune responses upon recognition of fungal cell wall chitin, followed by the release of various antimicrobials, including chitinase enzymes that hydrolyze chitin. In turn, many fungal pathogens secrete LysM effectors that prevent chitin recognition by the host through scavenging of chitin oligomers. We previously showed that intrachain LysM dimerization of the Cladosporium fulvum effector Ecp6 confers an ultrahigh-affinity binding groove that competitively sequesters chitin oligomers from host immune receptors. Additionally, particular LysM effectors are found to protect fungal hyphae against chitinase hydrolysis during host colonization. However, the molecular basis for the protection of fungal cell walls against hydrolysis remained unclear. Here, we determined a crystal structure of the single LysM domain-containing effector Mg1LysM of the wheat pathogen Zymoseptoria tritici and reveal that Mg1LysM is involved in the formation of two kinds of dimers; a chitin-dependent dimer as well as a chitin-independent homodimer. In this manner, Mg1LysM gains the capacity to form a supramolecular structure by chitin-induced oligomerization of chitin-independent Mg1LysM homodimers, a property that confers protection to fungal cell walls against host chitinases.

Klíčová slova:

Crystal structure – Dimerization – Fungal structure – Hydrolysis – Chitin – Plant cell walls – Plant fungal pathogens – Polymerization


1. Shibuya N, Kaku H, Kuchitsu K and Maliarik MJ. (1993) Identification of a novel high-affinity binding site for N-acetylchitooligosaccharide elicitor in the membrane fraction from suspension-cultured rice cells. FEBS Lett. 329: 75–78. doi: 10.1016/0014-5793(93)80197-3 8354412

2. Boller T. (2003) Chemoperception of microbial signals in plant cells. Annu. Rev. Plant Physiol. 46: 189–214.

3. Sanchez-Vallet A, Mesters JR and Thomma BPHJ. (2015) The battle for chitin recognition in plant-microbe interactions. FEMS Microbiol Rev. 39: 171–183. doi: 10.1093/femsre/fuu003 25725011

4. Schlumbaum A, Mauch F, Vögeli U and Boller T. (1986) Plant chitinases are potent inhibitors of fungal growth. Nature. 324: 365–367.

5. van Loon LC, Rep M and Pieterse CM. (2006) Significance of inducible defense-related proteins in infected plants. Annu Rev Phytopathol. 44: 135–162. doi: 10.1146/annurev.phyto.44.070505.143425 16602946

6. Adrangi S and Faramarzi MA. (2013) From bacteria to human: a journey into the world of chitinases. Biotechnol Adv. 31: 1786–1795. doi: 10.1016/j.biotechadv.2013.09.012 24095741

7. Hamid R, Khan MA, Ahmad M, Ahmad MM, Abdin MZ, Musarrat J, et al. (2013) Chitinases: An update. J Pharm Bioallied Sci. 5: 21–29. doi: 10.4103/0975-7406.106559 23559820

8. Kasprzewska A. (2003) Plant chitinases—regulation and function. Cell Mol Biol Lett. 8: 809–824. 12949620

9. Liu T, Chen L, Ma Q, Shen X and Yang Q. (2014) Structural insights into chitinolytic enzymes and inhibition mechanisms of selective inhibitors. Curr Pharm Des. 20: 754–770. doi: 10.2174/138161282005140214164730 23688083

10. Rovenich H, Zuccaro A and Thomma BP. (2016) Convergent evolution of filamentous microbes towards evasion of glycan-triggered immunity. New Phytol. 212: 896–901. doi: 10.1111/nph.14064 27329426

11. Kaku H, Nishizawa Y, Ishii-Minami N, Akimoto-Tomiyama C, Dohmae N, Takio K, et al. (2006) Plant cells recognize chitin fragments for defense signaling through a plasma membrane receptor. Proc Natl Acad Sci USA. 103: 11086–11091. doi: 10.1073/pnas.0508882103 16829581

12. Miya A, Albert P, Shinya T, Desaki Y, Ichimura K, Shirasu K, et al. (2007) CERK1, a LysM receptor kinase, is essential for chitin elicitor signaling in Arabidopsis. Proc Natl Acad Sci USA. 104: 19613–19618. doi: 10.1073/pnas.0705147104 18042724

13. Liu T, Liu Z, Song C, Hu Y, Han Z, She J, et al. (2012) Chitin-induced dimerization activates a plant immune receptor. Science. 336: 1160–1164. doi: 10.1126/science.1218867 22654057

14. de Jonge R, van Esse HP, Kombrink A, Shinya T, Desaki Y, Bours R, et al. (2010) Conserved fungal LysM effector Ecp6 prevents chitin-triggered immunity in plants. Science. 329: 953–955. doi: 10.1126/science.1190859 20724636

15. Marshall R, Kombrink A, Motteram J, Loza-Reyes E, Lucas J, Hammond-Kosack KE, et al. (2011) Analysis of two in planta expressed LysM effector homologs from the fungus Mycosphaerella graminicola reveals novel functional properties and varying contributions to virulence on wheat. Plant Physiol. 156: 756–769. doi: 10.1104/pp.111.176347 21467214

16. Okmen B, Kemmerich B, Hilbig D, Wemhoner R, Aschenbroich J, Perrar A, et al. (2018) Dual function of a secreted fungalysin metalloprotease in Ustilago maydis. New Phytol. 220: 249–261. doi: 10.1111/nph.15265 29916208

17. de Wit PJ. (2016) Cladosporium fulvum effectors: Weapons in the arms race with tomato. Annu Rev Phytopathol. 54: 1–23. doi: 10.1146/annurev-phyto-011516-040249 27215970

18. van den Burg HA, Harrison SJ, Joosten MH, Vervoort J and de Wit PJ. (2006) Cladosporium fulvum Avr4 protects fungal cell walls against hydrolysis by plant chitinases accumulating during infection. Mol Plant Microbe Interact. 19: 1420–1430. doi: 10.1094/MPMI-19-1420 17153926

19. van Esse HP, Bolton MD, Stergiopoulos I, de Wit PJ and Thomma BPHJ. (2007) The chitin-binding Cladosporium fulvum effector protein Avr4 is a virulence factor. Mol Plant Microbe Interact. 20: 1092–1101. doi: 10.1094/MPMI-20-9-1092 17849712

20. Sanchez-Vallet A, Saleem-Batcha R, Kombrink A, Hansen G, Valkenburg DJ, Thomma BPHJ, et al. (2013) Fungal effector Ecp6 outcompetes host immune receptor for chitin binding through intrachain LysM dimerization. Elife. 2: e00790. doi: 10.7554/eLife.00790 23840930

21. van den Burg HA, Spronk CA, Boeren S, Kennedy MA, Vissers JP, Vuister GW, et al. (2004) Binding of the AVR4 elicitor of Cladosporium fulvum to chitotriose units is facilitated by positive allosteric protein-protein interactions: the chitin-binding site of AVR4 represents a novel binding site on the folding scaffold shared between the invertebrate and the plant chitin-binding domain. J Biol Chem. 279: 16786–16796. doi: 10.1074/jbc.M312594200 14769793

22. Kohler AC, Chen LH, Hurlburt N, Salvucci A, Schwessinger B, Fisher AJ, et al. (2016) Structural analysis of an Avr4 effector ortholog offers insight into chitin binding and recognition by the Cf-4 receptor. Plant Cell. 28: 1945–1965. doi: 10.1105/tpc.15.00893 27401545

23. Hurlburt NK, Chen L-H, Stergiopoulos I and Fisher AJ. (2018) Structure of the Cladosporium fulvum Avr4 effector in complex with (GlcNAc) 6 reveals the ligand-binding mechanism and uncouples its intrinsic function from recognition by the Cf-4 resistance protein. PLoS Pathog. 14: e1007263. doi: 10.1371/journal.ppat.1007263 30148881

24. Stergiopoulos I, van den Burg HA, Okmen B, Beenen HG, van Liere S, Kema GH, et al. (2010) Tomato Cf resistance proteins mediate recognition of cognate homologous effectors from fungi pathogenic on dicots and monocots. Proc Natl Acad Sci USA. 107: 7610–7615. doi: 10.1073/pnas.1002910107 20368413

25. Bolton MD, van Esse HP, Vossen JH, de Jonge R, Stergiopoulos I, Stulemeijer IJ, et al. (2008) The novel Cladosporium fulvum lysin motif effector Ecp6 is a virulence factor with orthologues in other fungal species. Mol Microbiol. 69: 119–136. doi: 10.1111/j.1365-2958.2008.06270.x 18452583

26. de Jonge R and Thomma BPHJ. (2009) Fungal LysM effectors: extinguishers of host immunity? Trends Microbiol. 17: 151–157. doi: 10.1016/j.tim.2009.01.002 19299132

27. Kombrink A and Thomma BPHJ. (2013) LysM effectors: secreted proteins supporting fungal life. PLoS pathog. 9: e1003769. doi: 10.1371/journal.ppat.1003769 24348247

28. Kombrink A, Rovenich H, Shi-Kunne X, Rojas-Padilla E, van den Berg GC, Domazakis E, et al. (2017) Verticillium dahliae LysM effectors differentially contribute to virulence on plant hosts. Mol Plant Pathol. 18: 596–608. doi: 10.1111/mpp.12520 27911046

29. Mentlak TA, Kombrink A, Shinya T, Ryder LS, Otomo I, Saitoh H, et al. (2012) Effector-mediated suppression of chitin-triggered immunity by Magnaporthe oryzae is necessary for rice blast disease. Plant Cell. 24: 322–335. doi: 10.1105/tpc.111.092957 22267486

30. Takahara H, Hacquard S, Kombrink A, Hughes HB, Halder V, Robin GP, et al. (2016) Colletotrichum higginsianum extracellular LysM proteins play dual roles in appressorial function and suppression of chitin-triggered plant immunity. New Phytol. 211: 1323–1337. doi: 10.1111/nph.13994 27174033

31. Bergfors T. (2003) Seeds to crystals. J. Struct. Biol. 142: 66–76. doi: 10.1016/s1047-8477(03)00039-x 12718920

32. Bateman A and Bycroft M. (2000) The structure of a LysM domain from E. coli membrane-bound lytic murein transglycosylase D (MltD). J Mol Biol. 299: 1113–1119. doi: 10.1006/jmbi.2000.3778 10843862

33. Bielnicki J, Devedjiev Y, Derewenda U, Dauter Z, Joachimiak A and Derewenda ZS. (2006) B. subtilis ykuD protein at 2.0 A resolution: insights into the structure and function of a novel, ubiquitous family of bacterial enzymes. Proteins. 62: 144–151. doi: 10.1002/prot.20702 16287140

34. Koharudin LM, Viscomi AR, Montanini B, Kershaw MJ, Talbot NJ, Ottonello S, et al. (2011) Structure-function analysis of a CVNH-LysM lectin expressed during plant infection by the rice blast fungus Magnaporthe oryzae. Structure. 19: 662–674. doi: 10.1016/j.str.2011.03.004 21565701

35. Bozsoki Z, Cheng J, Feng F, Gysel K, Vinther M, Andersen KR, et al. (2017) Receptor-mediated chitin perception in legume roots is functionally separable from Nod factor perception. Proc Natl Acad Sci USA. 114: E8118–E8127. doi: 10.1073/pnas.1706795114 28874587

36. Krissinel E and Henrick K. (2007) Inference of macromolecular assemblies from crystalline state. J Mol Biol. 372: 774–797. doi: 10.1016/j.jmb.2007.05.022 17681537

37. Winn MD, Ballard CC, Cowtan KD, Dodson EJ, Emsley P, Evans PR, et al. (2011) Overview of the CCP4 suite and current developments. Acta Cryst. D. 67: 235–242.

38. Hartmann FE, Sanchez-Vallet A, McDonald BA and Croll D. (2017) A fungal wheat pathogen evolved host specialization by extensive chromosomal rearrangements. ISME J. 11: 1189–1204. doi: 10.1038/ismej.2016.196 28117833

39. Zeng T, Rodriguez-Moreno L, Mansurkhodzaev A, Wang P, van den Berg W, Gasciolli V, et al. (2020) A lysin motif effector subverts chitin-triggered immunity to facilitate arbuscular mycorrhizal symbiosis. New Phytol. 225: 448–460. doi: 10.1111/nph.16245 31596956

40. Newman J, Egan D, Walter TS, Meged R, Berry I, Ben Jelloul M, et al. (2005) Towards rationalization of crystallization screening for small- to medium-sized academic laboratories: the PACT/JCSG+ strategy. Acta Cryst. D. 61: 1426–1431.

41. Gerlach M, Mueller U and Weiss MS. (2016) The MX Beamlines BL14.1–3 at BESSY II. Journal of large-scale research facilities JLSRF. 2.

42. Adams PD, Afonine PV, Bunkoczi G, Chen VB, Davis IW, Echols N, et al. (2010) PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Cryst. D. 66: 213–221.

43. Murshudov GN, Skubak P, Lebedev AA, Pannu NS, Steiner RA, Nicholls RA, et al. (2011) REFMAC5 for the refinement of macromolecular crystal structures. Acta Cryst. D. 67: 355–367.

44. Emsley P, Lohkamp B, Scott WG and Cowtan K. (2010) Features and development of Coot. Acta Cryst. D. 66: 486–501.

45. Chen VB, Arendall WB 3rd, Headd JJ, Keedy DA, Immormino RM, Kapral GJ, et al. (2010) MolProbity: all-atom structure validation for macromolecular crystallography. Acta Cryst. D. 66: 12–21.

46. Hartmann FE and Croll D. (2017) Distinct trajectories of massive recent gene gains and losses in populations of a microbial eukaryotic pathogen. Mol Biol Evol. 34: 2808–2822. doi: 10.1093/molbev/msx208 28981698

47. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, et al. (2012) SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol. 19: 455–477. doi: 10.1089/cmb.2012.0021 22506599

48. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, et al. (2009) BLAST+: architecture and applications. BMC Bioinformatics. 10: 421. doi: 10.1186/1471-2105-10-421 20003500

49. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J, Homer N, et al. (2009) The Sequence Alignment/Map format and SAMtools. Bioinformatics. 25: 2078–2079. doi: 10.1093/bioinformatics/btp352 19505943

50. Katoh K and Standley DM. (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 30: 772–780. doi: 10.1093/molbev/mst010 23329690

51. Waterhouse AM, Procter JB, Martin DM, Clamp M and Barton GJ. (2009) Jalview Version 2—a multiple sequence alignment editor and analysis workbench. Bioinformatics. 25: 1189–1191. doi: 10.1093/bioinformatics/btp033 19151095

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