Molecular mapping with functional antibodies localizes critical sites on the human IL receptor common gamma (gamma c) chain.
The IL receptor common gamma (gamma c) chain is required for the formation of high affinity cytokine receptor complexes for IL-2, IL-4, IL-7, IL-9, and IL-15, and for signals regulating cell survival, growth, and differentiation. Our current understanding of how gamma c chain associates with multiple ligands and receptor subunits is drawn largely from its structural homology to the human growth hormone (hGH) receptor and known structure of the hGH/hGH receptor complex. These receptors share distinct features in their extracellular portions and are believed to function by a mechanism of ligand-induced association of receptor subunits. Here, we report the first directed mutational analysis of the human gamma c chain by alanine scanning conducted across seven regions likely to contain residues required for intermolecular contact. Functionally distinct, neutralizing anti-gamma c mAbs were employed to define critical residues. One particular mAb, CP.B8, unique in its ability to inhibit IL-2-, IL-4-, IL-7-, and IL-15-induced proliferation and high affinity cytokine binding of normal T cells as an intact mAb and as a Fab fragment, localized critical residues to four noncontinuous stretches, namely residues in loops AB and EF of domain 1, in the interdomain segment, and in loop FG of domain 2. Notably, these residues form a contiguous patch on the gamma c chain surface in a three-dimensional structural model. These results provide functional evidence for the location of contact points on gamma c chain required for its association with multiple ligands.
Biogen, Inc., Cambridge, MA 02142, USA.
N
N
Raskin
A
A
Jakubowski
I D
ID
Sizing
D L
DL
Olson
S L
SL
Kalled
C A
CA
Hession
C D
CD
Benjamin
D P
DP
Baker
L C
LC
Burkly
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D000918:Antibody Specificity / Q000235:genetics; D001666:Binding Sites, Antibody / Q000235:genetics; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000276:immunology; D005260:Female; D006801:Humans; D051379:Mice; D008817:Mice, Mutant Strains; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018121:Receptors, Cytokine / Q000276:immunology; D018123:Receptors, Interleukin / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D016207:Cytokines; D018121:Receptors, Cytokine; D018123:Receptors, Interleukin; D000409:Alanine