Ligation of SIPRa on macrophages by CD47 on RBCs inhibits FcgR-mediated phagocytosis by promoting tyrosine phosphorylation of SIPRa and its interaction with SHP-1 and preventing SIPRa

crucial for tissue homeostasis, the resolution of inflammation, the modulation of immune responses and cancer immunotherapy. Apoptosis is a dynamic process, during which cell surface molecules are continuously changing. Enhanced ‘eat me’ signals and reduced ‘don’t eat me’ signals in apoptotic cells induce phagocytes to promptly and efficiently engulf apoptotic cells during the early and late stages of cell death, preventing them from releasing noxious intracellular contents. Direct ‘eat me’ signals include the newly expressed molecules phosphatidylserine (PS) and annexin I, modification of the surface molecules intercellular cell adhesion molecule (ICAM)-3 and platelet endothelial cell adhesion molecule-1 (PECAM-1, CD31), changes in the cell surface charge, and presence of extracellular bridging molecules such as milk-fat-globule-EGF-factor 8, the collectin family proteins, mannose-binding protein, complement C1q, and thrombospondin-1. However, very little is known about changes in the ‘don’t eat me’ signals during apoptosis. CD47 (integrin-associated protein, IAP) is a 50 kDa integral membrane protein composed of an extracellular immunoglobulin variable (IgV) domain, five transmembrane domains, and a short C-terminal cytoplasmic tail with four alternatively spliced forms, and is a widely expressed member of the Ig superfamily. CD47 functions as a receptor for thrombospondin-1 and as a ligand for the transmembrane signal regulatory proteins (SIRP)-a and -g , and also interacts with integrins such as the RGD receptor avb3, the fibrinogen receptor a llbb3, and the collagen receptor a2b1 to induce various cellular signaling. Integrin avb3/CD47 ligation initiates a signal transduction cascade, dependent upon the b3 cytoplasmic tail, which inhibits the phagocytic function of a5b1 at a step subsequent to modulation of integrin affinity. CD47 on red blood cells (RBCs) functions as a ‘don’t eat me’ marker by sending a negative engulfment signal to splenic red pulp macrophages through SIPRa . Most signaling through CD47-SIPRa interaction is mediated by the phosphatase Src homology 2 domain-containing protein tyrosine phosphatase (SHP-1). Ligation of SIPRa on macrophages by CD47 on RBCs inhibits FcgR-mediated phagocytosis by promoting tyrosine phosphorylation of SIPRa and its interaction with SHP-1 and preventing SIPRa dephosphorylation in response to FcgR activation. CD47 is altered and/or lost on apoptotic cells, and disruption of interactions between CD47 on apoptotic cells and SIPRa on the engulfing cells permits uptake of apoptotic cells in a calreticulin/LDL receptor-related protein (LRP)-dependent manner. Phagocytosis of opsonized RBCs by splenic macrophages from mice that express the mutant form of SIPRa (lacking most of its cytoplasmic region) is increased compared with wild-type macrophages. However, macrophages require CD47 and PS on apoptotic cells for engulfment and CD47-SIPRa interaction works as a tethering step in the phagocytosis in lymphocytes. CD47 on healthy cells and its engagement of the phagocytic receptor SIPRa appears to contribute a key ‘don’t eat me’ signal. CD31 is a 130 kDa glycoprotein belonging to the Ig superfamily of cell adhesion molecules and is restrictedly expressed in the vascular system, platelets, monocytes, neutrophils, selected T-cells and endothelial cells. CD31 is composed of six extracellular Ig-domains, comprising homophilic interaction involving Ig domain 1 and heterophilic ligands integrin a vb 3 and CD38 involving Ig-domains 1—3. CD31 signaling is mediated through phosphorylation of specific tyrosine residues located in two immunoregulatory tyrosine-based inhibition motif (ITIM) domains in its cytoplasmic tail and interactions with SHP-2 and SHP-1. CD31 homophilic interaction induces cytoprotective signaling to inhibit Bax-mediated mitochondrial damage by SHP-2 inter1828 Vol. 34, No. 12 Regular Article