The major goal of this proposal is to dissect, by molecular and biochemical approaches, the events involved in phagocytosis. This complex cellular event is a specialized feature of monocytes, macrophages of polymorpholeukocytes. Three major phagocytic receptors have been defined on the surface of phagocytes. The receptors for the Fc portion of immunoglobulin and complement require the presence of their respective opsonins in order to recognize targets. The mannose receptor, however, recognizes various configurations of mannose, N-acetylglyglucosamine and other sugars that are ubiquitously expressed on the surface of pathogens. Removal of red blood cells, platelets, and neutrophils opsonized by autoreactive opsonins occurs via phagocytic receptors expressed on tissue macrophages in the spleen and liver as well as other sites of the reticular endothelial system including the bone marrow. Although it is well established that receptor ligand interactions occur sequentially around the full circumference of the target, like a zipper, very little is known about the initial biochemical events that are involved in phagocytosis. Previously, it was thought that phagocytosis was dependent upon the expression of a host of myeloid specific genes whose products interacted to mediate this complex cellular event. Recent experiments from our laboratory have challenged this idea as we were able to demonstrate that transient expression of Cos-I epithelial cells with the macrophage mannose receptor cDNA enabled these cells to phagocytose particles. Mutant receptors lacking the mannose receptor tail are expressed on the cell surface but are unable to ingest bound particles. These. experiments suggested that ligation of the ectodomain of the mannose receptor by a multivalent ligand was able to signal phagocytosis that was cytochalasin sensitive in a heterologous cell. Work from other laboratories indicated that some subclasses of Fc receptors, FcRII, were able to phagocytose opsonized heat killed toxoplasma in Chinese hamster ovary cells whilst another FcR, Fc-lambda R1, was able to bind but not internalize opsonized erythrocytes when transfected into Cos cells. We have been able to create a 'gain of function' chimeric receptor by transferring the mannose receptor transmembrane region and cytoplasmic tail to the ectodomain of Fc-lambda R1. Equivalent rosetting of opsonized erythrocytes is observed when 'wild type' Fc-lambda R1 and Fc-lambda R1 ectodomain MR transmembrane region and MR tail are transfected into Cos cells but only the latter receptors are able to mediate ingestion of bound particles. These 'gain of function' chimeric receptors indicate that the MR transmembrane region and cytoplasmic tail are able to engage cellular machinery in non-myeloid cells and induce phagocytosis. The two primary goals are (1) to define distinct motif(s) that are critical for phagocytosis; (2) characterize the cellular proteins that interact with the mannose receptor cytoplasmic tail with a view to discerning the early biochemical events of phagocytosis.
| Eichbaum, Q; Heney, D; Raveh, D et al. (1997) Murine macrophage mannose receptor promoter is regulated by the transcription factors PU.1 and SP1. Blood 90:4135-43 |
