title: Two modes of beta-receptor recognition are mediated by distinct epitopes on mouse and human interleukin-3.
abstract: The cytokine interleukin-3 (IL-3) is a critical regulator of inflammation and immune responses in mammals. IL-3 exerts its effects on target cells via receptors comprising an IL-3-specific alpha-subunit and common beta-subunit (beta c; shared with IL-5 and granulocyte-macrophage colony-stimulating factor) or a beta-subunit that specifically binds IL-3 (beta(IL-3); present in mice but not humans). We recently identified two splice variants of the alpha-subunit of the IL-3 receptor (IL-3R alpha) that are relevant to hematopoietic progenitor cell differentiation or proliferation: the full length ("SP1" isoform) and a novel isoform (denoted "SP2") lacking the N-terminal Ig-like domain. Although our studies demonstrated that each mouse IL-3 (mIL-3) R alpha isoform can direct mIL-3 binding to two distinct sites on the beta(IL-3) subunit, it has remained unclear which residues in mIL-3 itself are critical to the two modes of beta(IL-3) recognition and whether the human IL-3R alpha SP1 and SP2 orthologs similarly instruct human IL-3 binding to two distinct sites on the human beta c subunit. Herein, we describe the identification of residues clustering around the highly conserved A-helix residue, Glu(23), in the mIL-3 A- and C-helices as critical for receptor binding and growth stimulation via the beta(IL-3) and mIL-3R alpha SP2 subunits, whereas an overlapping cluster was required for binding and activation of beta(IL-3) in the presence of mIL-3R alpha SP1. Similarly, our studies of human IL-3 indicate that two different modes of beta c binding are utilized in the presence of the hIL-3R alpha SP1 or SP2 isoforms, suggesting a possible conserved mechanism by which the relative orientations of receptor subunits are modulated to achieve distinct signaling outcomes.
authors.affiliation: Department of Structural Biology, John Curtin School of Medical Research, Australian National University, Canberra, Australian Capital Territory 0200, Australia.
authors.forename: Shamaruh
authors.initials: S
authors.lastname: Mirza
authors.forename: Jinglong
authors.initials: J
authors.lastname: Chen
authors.forename: Bin
authors.initials: B
authors.lastname: Wen
authors.forename: Cameron L
authors.initials: CL
authors.lastname: Ewens
authors.forename: Jin
authors.initials: J
authors.lastname: Dai
authors.forename: James M
authors.initials: JM
authors.lastname: Murphy
authors.forename: Ian G
authors.initials: IG
authors.lastname: Young
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D049109:Cell Proliferation; D053648:Cytokine Receptor Common beta Subunit / Q000737:chemistry / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004789:Enzyme Activation; D000939:Epitopes / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D006801:Humans; D007377:Interleukin-3 / Q000737:chemistry / Q000378:metabolism*; D053650:Interleukin-3 Receptor alpha Subunit / Q000378:metabolism; D053614:Janus Kinase 2 / Q000378:metabolism; D020935:MAP Kinase Signaling System; D051379:Mice; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism
chemical_list: D053648:Cytokine Receptor Common beta Subunit; D000939:Epitopes; C508596:IL3 protein, human; D007377:Interleukin-3; D053650:Interleukin-3 Receptor alpha Subunit; D050505:Mutant Proteins; D020033:Protein Isoforms; D018698:Glutamic Acid; D053614:Janus Kinase 2
references.citation: J Biol Chem. 1995 Oct 6;270(40):23754-60
references.pmid: 7559548
references.citation: Growth Factors. 2010 Apr;28(2):104-10
references.pmid: 19961363
references.citation: Biomol NMR Assign. 2010 Apr;4(1):73-7
references.pmid: 20174897
references.citation: Vitam Horm. 2006;74:1-30
references.pmid: 17027509
references.citation: Nat Struct Mol Biol. 2005 Sep;12(9):814-21
references.pmid: 16116438
references.citation: EMBO J. 1995 Jul 17;14(14):3395-402
references.pmid: 7628440
references.citation: J Biol Chem. 2009 Feb 27;284(9):5763-73
references.pmid: 19109256
references.citation: J Biol Chem. 1996 Oct 18;271(42):25999-6006
references.pmid: 8824238
references.citation: J Biol Chem. 1996 Dec 13;271(50):31922-8
references.pmid: 8943237
references.citation: J Exp Med. 1993 Jun 1;177(6):1523-9
references.pmid: 8496674
references.citation: Eur J Biochem. 1999 Jul;263(2):438-46
references.pmid: 10406952
references.citation: Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32
references.pmid: 15572765
references.citation: Nat Immunol. 2009 Jul;10(7):697-705
references.pmid: 19465906
references.citation: Mol Cell Biol. 1994 Jul;14(7):4335-41
references.pmid: 8007942
references.citation: J Pharmacol Methods. 1985 Nov;14(3):213-28
references.pmid: 3840547
references.citation: EMBO J. 1992 May;11(5):1875-84
references.pmid: 1582416
references.citation: J Leukoc Biol. 2007 Apr;81(4):1137-48
references.pmid: 17227823
references.citation: Dev Cell. 2006 Aug;11(2):171-80
references.pmid: 16890157
references.citation: J Biol Chem. 1994 Mar 18;269(11):8488-92
references.pmid: 7907592
references.citation: EMBO J. 1989 Dec 1;8(12):3667-76
references.pmid: 2555171
references.citation: Nat Immunol. 2009 Jul;10(7):706-12
references.pmid: 19465908
references.citation: Proc Natl Acad Sci U S A. 1990 Dec;87(24):9655-9
references.pmid: 1702217
references.citation: Cell. 2001 Jan 26;104(2):291-300
references.pmid: 11207369
references.citation: Biochem J. 2010 Feb 24;426(3):307-17
references.pmid: 20078425
references.citation: EMBO J. 1992 Oct;11(10):3541-9
references.pmid: 1396555
references.citation: Science. 1990 Jan 19;247(4940):324-7
references.pmid: 2404337
references.citation: Mol Cell. 2003 Nov;12(5):1239-50
references.pmid: 14636581
references.citation: Cell. 2008 Aug 8;134(3):496-507
references.pmid: 18692472
references.citation: J Biol Chem. 2003 Mar 21;278(12):10572-7
references.pmid: 12525483
references.citation: Anal Biochem. 1980 Sep 1;107(1):220-39
references.pmid: 6254391
references.citation: Cell. 1991 Sep 20;66(6):1175-84
references.pmid: 1833065
references.citation: Nat Immunol. 2009 Jul;10(7):713-20
references.pmid: 19465907
references.citation: Cytokine. 2008 May;42(2):234-242
references.pmid: 18374598
references.citation: Proc Natl Acad Sci U S A. 1995 May 23;92(11):5194-8
references.pmid: 7761472
references.citation: Nature. 1998 Mar 5;392(6671):90-3
references.pmid: 9510253
references.citation: Cell. 1991 Sep 20;66(6):1165-74
references.pmid: 1833064
references.citation: J Biol Chem. 1995 Jun 30;270(26):15762-9
references.pmid: 7797578
references.citation: J Biol Chem. 2004 Jun 18;279(25):26500-8
references.pmid: 15060062
references.citation: Eur J Biochem. 2001 May;268(10):2905-11
references.pmid: 11358507
references.citation: EMBO J. 1994 Nov 1;13(21):5176-85
references.pmid: 7957082
references.citation: EMBO J. 1992 Mar;11(3):909-16
references.pmid: 1532144
references.citation: Nat Methods. 2006 Apr;3(4):287-93
references.pmid: 16554834