Molecular dissection of human interleukin-31-mediated signal transduction through site-directed mutagenesis.
Interleukin (IL)-31 is a recently described cytokine, preferentially produced by T helper 2 lymphocytes and associated with skin diseases, such as atopic dermatitis. IL-31 is a member of the four alpha-helix bundle cytokine family and is related to the IL-6 subgroup. Its heterodimeric membrane receptor is composed of the gp130-like receptor (GPL) subunit associated to the oncostatin M receptor subunit. We identified critical amino acids implicated in the ligand receptor interaction by computational analysis combined with site-directed mutagenesis. Six IL-31 residues selected for their putative involvement in cytokine receptor contact sites were alanine-substituted, and the corresponding proteins were expressed in mammalian and bacterial systems. Biochemical, membrane binding, cell signaling, and cell proliferation analyses showed that mutation E44A, E106A, or H110A abolished IL-31 binding to GPL and the subsequent signaling events. A second ligand receptor-binding site involved Lys(134), with alanine substitution leading to a protein that still binds GPL, but is unable to recruit the second receptor subunit and the subsequent signaling pathways. The results indicate that IL-31 recognizes its receptor complex through two different binding sites, and we propose a three-dimensional model for IL-31.
Le Saux
Sabine
S
Unité Mixte INSERM 564, Bâtiment Monteclair, 4 rue Larrey, 49933 Angers Cedex 09, France.
Rousseau
François
F
Barbier
Fabien
F
Ravon
Elisa
E
Grimaud
Linda
L
Danger
Yannic
Y
Froger
Josy
J
Chevalier
Sylvie
S
Gascan
Hugues
H
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D049109:Cell Proliferation; D002522:Chlorocebus aethiops; D006801:Humans; D007378:Interleukins / Q000235:genetics* / Q000378:metabolism*; D008024:Ligands; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D053677:Receptors, Oncostatin M / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction
D000596:Amino Acids; C502187:IL31 protein, human; D007378:Interleukins; D008024:Ligands; D053677:Receptors, Oncostatin M