Receptor modulation by Fc gamma RI-specific fusion proteins is dependent on receptor number and modified by IgG.
The high-affinity IgG receptor, FcgammaRI (CD64), is constitutively expressed exclusively on professional APCs. Human FcgammaRI binds monomeric IgG with high affinity and is, therefore, saturated in vivo. The binding of IgG to FcgammaRI causes receptor recycling, while Abs that cross-link FcgammaRI cause rapid down-modulation of surface FcgammaRI. Because studies performed in the absence of ligand may not be representative of FcgammaRI modulation in vivo, we investigated the ability of FcgammaRI-cross-linking Abs and non-cross-linking derivatives to modulate FcgammaRI in the presence and absence of ligand. In the absence of ligand mAb H22 and wH22xeGFP, an enhanced green fluorescent protein (eGFP)-labeled fusion protein of H22, cross-linked and rapidly down-modulated surface FcgammaRI on the human myeloid cell line, U937, and its high FcgammaRI-expressing subclone, 10.6. This effect was dependent on the concentration of fusion protein and the level of FcgammaRI expression and correlated with internalization of both wH22xeGFP and FcgammaRI, itself, as assessed by confocal microscopy. A single-chain Fv version, sFv22xeGFP, which does not cross-link FcgammaRI, was unable to modulate FcgammaRI in the absence of IgG. However, if ligand was present, treatment with either monovalent or cross-linking fusion protein led to intracellular receptor accumulation. These findings suggest at least two alternate mechanisms of internalization that are influenced by ligand and demonstrate the physiologic potential of FcgammaRI to transport a large antigenic load into APCs for processing. These studies may lead to the development of better FcgammaRI-targeted vaccines, as well as therapies to down-modulate FcR involved in autoimmune diseases.
Guyre
C A
CA
Department of Physiology, Dartmouth Medical School, Lebanon, NH 03756, USA.
Keler
T
T
Swink
S L
SL
Vitale
L A
LA
Graziano
R F
RF
Fanger
M W
MW
D000276:Adjuvants, Immunologic / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D001666:Binding Sites, Antibody / Q000235:genetics; D002999:Clone Cells; D004306:Dose-Response Relationship, Immunologic; D015536:Down-Regulation / Q000235:genetics / Q000276:immunology; D005822:Genetic Vectors / Q000378:metabolism / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology*; D007424:Intracellular Fluid / Q000276:immunology / Q000378:metabolism; D008024:Ligands; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018448:Models, Immunological; D017452:Receptors, IgG / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000276:immunology* / Q000378:metabolism* / Q000494:pharmacology; D020298:U937 Cells
D000276:Adjuvants, Immunologic; D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D008024:Ligands; D008164:Luminescent Proteins; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins