Abstract
Clustering of macrophage Fc gamma receptors by multimeric immunoglobulin complexes leads to their internalization. Formation of small aggregates leads to endocytosis, whereas large particulate complexes induce phagocytosis. In RAW-264.7 macrophages, Fc gamma receptor endocytosis was found to be dependent on clathrin and dynamin and insensitive to cytochalasin. Clathrin also associates with nascent phagosomes, and earlier observations suggested that it plays an essential role in phagosome formation. However, we find that phagocytosis of IgG-coated large (> or =3 microm) particles was unaffected by inhibition of dynamin or by reducing the expression of clathrin using antisense mRNA but was eliminated by cytochalasin, implying a distinct mechanism dependent on actin assembly. The uptake of smaller particles (< or =1 microm) was only partially blocked by cytochalasin. Remarkably, the cytochalasin-resistant component was also insensitive to dominant-negative dynamin I and to clathrin antisense mRNA, implying the existence of a third internalization mechanism, independent of actin, dynamin, and clathrin. The uptake of small particles occurred by a process distinct from fluid phase pinocytosis, because it was not inhibited by dominant-negative Rab5. The insensitivity of phagocytosis to dominant-negative dynamin I enabled us to test the role of dynamin in phagosomal maturation. Although internalization of receptors from the plasma membrane was virtually eliminated by the K44A and S45N mutants of dynamin I, clearance of transferrin receptors and of CD18 from maturing phagosomes was unaffected by these mutants. This implies that removal of receptors from the phagosomal membrane occurs by a mechanism that is different from the one mediating internalization of the same receptors at the plasma membrane. These results imply that, contrary to prevailing notions, normal dynamin and clathrin function is not required for phagocytosis and reveal the existence of a component of phagocytosis that is independent of actin and Rab5.
Highlights
Receptors on the surface of phagocytes are essential in the recognition and elimination of microorganisms
Upon binding to Fcγ-receptors, immunoglobulin G (IgG)-containing immune complexes are internalized and travel along the endocytic pathway where they are targeted for degradation
Transient expression of a fusion of the light chain of clathrin with EGFP was used to examine whether clathrin is involved in the receptor-mediated endocytosis of aggregated IgG by RAW 264.7 cells
Summary
Receptors on the surface of phagocytes are essential in the recognition and elimination of microorganisms. Some phagocytic receptors interact directly with the pathogens, by recognizing motifs that are conserved on the surface of various types of microorganisms [1,2]. A second group of receptors interact instead with opsonins, host molecules such as antibodies or complement components that coat the surface of the microorganisms [3]. The latter class includes Fcγ receptors, which bind to the constant region of immunoglobulin G (IgG). Abnormal function or regulation of Fcγ receptors may lead to the ineffective clearance of infectious organisms or can cause immunohematologic and autoimmune disorders [4]
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