Phagocytosis in macrophages lacking Cbl reveals an unsuspected role for Fc gamma receptor signaling and actin assembly in target binding.
Fc gamma receptor (Fc gammaR)-mediated phagocytosis is known to require tyrosine kinases (TKs). We identified c-Cbl and Cbl-b as proteins that undergo tyrosine phosphorylation during phagocytosis. Cbl-deficient macrophages displayed enhanced Fc gammaR-mediated signaling and phagocytosis. Surprisingly, binding of IgG-coated targets (EIgG) was also enhanced. c-Cbl-deficient macrophages expressed less Fc gammaRIIb, the inhibitory Fc gamma receptor; however, this did not account for enhanced target binding. We isolated the function of one Fc receptor isoform, Fc gammaRI, using IgG2a-coated targets (EIgG2a). Cbl-deficient macrophages demonstrated a disproportionate increase in binding EIgG2a, suggesting that signal strength regulates binding efficiency toward opsonized targets. In resting cells, Fc gammaRI colocalized with the Src family TK Hck in F-actin-rich structures, which was enhanced in Cbl-deficient macrophages. Target binding was sensitive to TK inhibitors, profoundly inhibited following depletion of cholesterol, and ablated at 4 degrees C or in the presence of inhibitors of actin polymerization. Sensitivity of EIgG binding to cytoskeletal disruption was inversely proportional to opsonin density. These findings challenge the view that Fc gammaR-mediated binding is a passive event. They suggest that dynamic engagement of TKs and the cytoskeleton enables macrophages to serve as cellular "Venus fly traps", with the capacity to capture phagocytic targets under conditions of limiting opsonin density.
Dale
Benjamin M
BM
Department of Pharmacology, Columbia University College of Physicians & Surgeons, New York, NY 10032, USA.
Traum
Daniel
D
Erdjument-Bromage
Hediye
H
Tempst
Paul
P
Greenberg
Steven
S
D000199:Actins / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D001854:Bone Marrow Cells / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D015536:Down-Regulation / Q000276:immunology; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010587:Phagocytosis / Q000276:immunology*; D010766:Phosphorylation; D011485:Protein Binding / Q000276:immunology; D050721:Proto-Oncogene Proteins c-cbl / Q000172:deficiency* / Q000235:genetics* / Q000502:physiology; D017452:Receptors, IgG / Q000037:antagonists & inhibitors / Q000502:physiology*; D015398:Signal Transduction / Q000276:immunology*; D014443:Tyrosine / Q000378:metabolism
D000199:Actins; D017452:Receptors, IgG; D014443:Tyrosine; D050721:Proto-Oncogene Proteins c-cbl; C493849:Cbl protein, mouse