Receptor modulation by Fc gamma RI-specific fusion proteins is dependent on receptor number and modified by IgG

The high-affinity IgG receptor, Fc gamma RI (CD64), is constitutively expre ssed exclusively on professional APCs. Human Fc gamma RI binds monomeric Ig G with high affinity and is, therefore, saturated in vivo. The binding of I gG to Fc gamma RI causes receptor recycling, while Abs that cross-link Fc g amma RI cause rapid down-modulation of surface Fc gamma RI. Because studies performed in the absence of ligand may not be representative of Fc gamma R I modulation in vivo, we investigated the ability of Fc gamma RI-cross-link ing Abs and non-cross-linking derivatives to modulate Fc gamma RI in the pr esence and absence of ligand. In the absence of ligand mAb H22 and wH22xeGF P, an enhanced green fluorescent protein (eGFP)-labeled fusion protein of H 22, cross-linked and rapidly down-modulated surface Fc gamma RI on the huma n myeloid cell line, U937, and its high Fc gamma RI-expressing subclone, 10 .6. This effect was dependent on the concentration of fusion protein and th e level of Fc gamma RI expression and correlated with internalization of bo th wH22xeGFP and Fc gamma RI, itself, as assessed by confocal microscopy. A single-chain Fv version, sFv22xeGFP, which does not cross-link Fc gamma RI , was unable to modulate Fc gamma RI in the absence of IgG. However, if lig and was present, treatment with either monovalent or crosslinking fusion pr otein led to intracellular receptor accumulation. These findings suggest at least two alternate mechanisms of internalization that are influenced by l igand and demonstrate the physiologic potential of Fc gamma RI to transport a large antigenic load into APCs for processing. These studies may lead to the development of better Fc gamma RI-targeted vaccines, as well as therap ies to down-modulate FcR involved in autoimmune diseases.