title: Dimerization of the fungal defense lectin CCL2 is essential for its toxicity against nematodes.
abstract: Lectins are used as defense effector proteins against predators, parasites and pathogens by animal, plant and fungal innate defense systems. These proteins bind to specific glycoepitopes on the cell surfaces and thereby interfere with the proper cellular functions of the various antagonists. The exact cellular toxicity mechanism is in many cases unclear. Lectin CCL2 of the mushroom Coprinopsis cinerea was previously shown to be toxic for Caenorhabditis elegans and Drosophila melanogaster. This toxicity is dependent on a single, high-affinity binding site for the trisaccharide GlcNAc(Fucα1,3)β1,4GlcNAc, which is a hallmark of nematode and insect N-glycan cores. The carbohydrate-binding site is located at an unusual position on the protein surface when compared to other β-trefoil lectins. Here, we show that CCL2 forms a compact dimer in solution and in crystals. Substitution of two amino acid residues at the dimer interface, R18A and F133A, interfered with dimerization of CCL2 and reduced toxicity but left carbohydrate-binding unaffected. These results, together with the positioning of the two carbohydrate-binding sites on the surface of the protein dimer, suggest that crosslinking of N-glycoproteins on the surface of intestinal cells of invertebrates is a crucial step in the mechanism of CCL2-mediated toxicity. Comparisons of the number and positioning of carbohydrate-binding sites among different dimerizing fungal β-trefoil lectins revealed a considerable variability in the carbohydrate-binding patterns of these proteins, which are likely to correlate with their respective functions.
authors.affiliation: Institute of Microbiology, ETH Zürich, Vladimir-Prelog-Weg 4, 8093 Zürich, Switzerland.
authors.forename: Silvia
authors.initials: S
authors.lastname: Bleuler-Martinez
authors.affiliation: Institute of Molecular Life Sciences, University of Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland.
authors.forename: Katrin
authors.initials: K
authors.lastname: Stutz
authors.affiliation: Institute of Microbiology, ETH Zürich, Vladimir-Prelog-Weg 4, 8093 Zürich, Switzerland.
authors.forename: Ramon
authors.initials: R
authors.lastname: Sieber
authors.affiliation: Laboratoire des Biomolécules, UPMC Université Paris 06, Ecole Normale Supérieure, Paris, France.
authors.forename: Mayeul
authors.initials: M
authors.lastname: Collot
authors.affiliation: Laboratoire des Biomolécules, UPMC Université Paris 06, Ecole Normale Supérieure, Paris, France.
authors.forename: Jean-Maurice
authors.initials: JM
authors.lastname: Mallet
authors.affiliation: Institute of Molecular Life Sciences, University of Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland.
authors.forename: Michael
authors.initials: M
authors.lastname: Hengartner
authors.affiliation: Institute of Molecular Biology and Biophysics, ETH Zürich, Schafmattstr. 20, 8093 Zürich, Switzerland.|Department of Molecular Biology, University of Salzburg, Billrothstrasse 11, 5020 Salzburg, Austria.
authors.forename: Mario
authors.initials: M
authors.lastname: Schubert
authors.affiliation: CERMAV, UPR5301, CNRS and Université Grenoble Alpes, 38041 Grenoble, France.
authors.forename: Annabelle
authors.initials: A
authors.lastname: Varrot
authors.affiliation: Institute of Microbiology, ETH Zürich, Vladimir-Prelog-Weg 4, 8093 Zürich, Switzerland.
authors.forename: Markus
authors.initials: M
authors.lastname: Künzler
mesh_terms: D000363:Agaricales / Q000737:chemistry*; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D017173:Caenorhabditis elegans / Q000472:pathogenicity; D019281:Dimerization; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism; D037181:Lectins, C-Type / Q000737:chemistry* / Q000378:metabolism; D011134:Polysaccharides / Q000235:genetics / Q000378:metabolism; D014312:Trisaccharides / Q000737:chemistry* / Q000235:genetics
chemical_list: D005656:Fungal Proteins; D037181:Lectins, C-Type; D011134:Polysaccharides; D014312:Trisaccharides
keywords: carbohydrate recognition; fucosylated N-glycan core; lectin dimer; nematodes; toxin