Two distinct cytoplasmic regions of the β₂ integrin chain regulate RhoA function during phagocytosis

α_Mβ₂ integrins mediate phagocytosis of opsonized particles in a process controlled by RhoA, Rho kinase, myosin II, Arp2/3, and actin polymerization. α_Mβ₂, Rho, Arp2/3, and F-actin accumulate underneath bound particles; however, the mechanism regulating Rho function during α_Mβ₂-mediated phagocytosis is poorly understood. We report that the binding of C3bi-opsonized sheep red blood cells (RBCs) to α_Mβ₂ increases Rho-GTP, but not Rac-GTP, levels. Deletion of the cytoplasmic domain of β₂, but not of α_M, abolished Rho recruitment and activation, as well as phagocytic uptake. Interestingly, a 16–amino acid (aa) region in the membrane-proximal half of the β₂ cytoplasmic domain was necessary for activating Rho. Three COOH-terminal residues (aa 758–760) were essential for β₂-induced accumulation of Rho at complement receptor 3 (CR3) phagosomes. Activation of Rho was necessary, but not sufficient, for its stable recruitment underneath bound particles or for uptake. However, recruitment of active Rho was sufficient for phagocytosis. Our data shed light on the mechanism of outside-in signaling, from ligated integrins to the activation of Rho GTPase signaling.