title: A functional interleukin 12 receptor complex is composed of two beta-type cytokine receptor subunits.
abstract: We have identified a cDNA from a human phytohemagglutinin-activated lymphoblast library encoding a protein that binds 125I-labeled human interleukin 12 (125I-huIL-12) with a Kd of about 5 nM when expressed in COS-7 cells. When coexpressed in COS-7 cells with the previously identified IL-12 beta receptor (IL-12R beta) protein, two classes of 125I-huIL-12 binding sites were measured with Kds of about 55 pM and 8 nM, corresponding to the high- and low-affinity binding sites seen on phytohemagglutinin-activated lymphoblasts. This newly identified huIL-12R subunit is a member of the cytokine receptor superfamily, with closest resemblance to the beta-type cytokine receptor gp130 and the receptors for leukemia inhibitory factor and granulocyte colony-stimulating factor. Consequently, we have reclassified the previously identified IL-12R beta subunit as huIL-12R beta 1 and designated the newly identified subunit as huIL-12R beta 2. huIL-12R beta 2 is an 862-amino acid type I transmembrane protein with a 595-amino-acid-long extracellular domain and a cytoplasmic tail of 216 amino acids that contains three tyrosine residues. A cDNA encoding the mouse homolog of the huIL12R beta 2 protein has also been isolated. Human and mouse IL-12R beta 2 proteins show a 68% amino acid sequence identity. When expressed in COS-7 cells, huIL-12R beta 2 exists as a disulfide-linked oligomer with an apparent monomeric molecular weight of 130 kDa. Coexpression of the two identified IL-12R subunits in Ba/F3 cells conferred IL-12 responsiveness, and clones of these cotransfected Ba/F3 cells that grew continuously in the presence of IL-12 were isolated and designated LJM-1 cells. LJM-1 cells exhibited dose-dependent proliferation in response to huIL-12, with an ED50 of about 1 pM huIL-12. Interestingly, Ba/F3 cells transfected with IL-12R beta 2 alone proliferated in response to huIL-12 with an ED50 of about 50 pM, although a role for endogenous mouse IL-12R beta 1 in IL-12 signal transduction in these transfectants cannot be ruled out. These results demonstrate that the functional high-affinity IL-12R is composed of at least two beta-type cytokine receptor subunits, each independently exhibiting a low affinity for IL-12.
authors.affiliation: Department of Inflammation/Autoimmune Diseases, Hoffmann-La Roche, Inc., Nutley, NJ 07110, USA.
authors.forename: D H
authors.initials: DH
authors.lastname: Presky
authors.forename: H
authors.initials: H
authors.lastname: Yang
authors.forename: L J
authors.initials: LJ
authors.lastname: Minetti
authors.forename: A O
authors.initials: AO
authors.lastname: Chua
authors.forename: N
authors.initials: N
authors.lastname: Nabavi
authors.forename: C Y
authors.initials: CY
authors.lastname: Wu
authors.forename: M K
authors.initials: MK
authors.lastname: Gately
authors.forename: U
authors.initials: U
authors.lastname: Gubler
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002462:Cell Membrane / Q000276:immunology; D002478:Cells, Cultured; D003001:Cloning, Molecular; D006801:Humans; D018664:Interleukin-12 / Q000378:metabolism; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D018121:Receptors, Cytokine / Q000096:biosynthesis* / Q000737:chemistry; D018123:Receptors, Interleukin / Q000096:biosynthesis* / Q000737:chemistry; D053707:Receptors, Interleukin-12; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014162:Transfection
chemical_list: D046911:Macromolecular Substances; D018121:Receptors, Cytokine; D018123:Receptors, Interleukin; D053707:Receptors, Interleukin-12; D011994:Recombinant Proteins; D018664:Interleukin-12