Supplementary MaterialsSupplementary Shape 1. tethering receptor without a putative co-receptor. Phagocytes expressing Anxa5-GPI exhibited enhanced binding of apoptotic cells, resulting in promoted ingestion of apoptotic cells in a phosphatidylserine-dependent manner. Anxa5-GPI-induced phagocytosis of apoptotic cells relied on the known cytoskeletal engulfment machinery but partially depended on the Elmo-Dock-Rac module or the integrin pathway. In addition, Anxa5-GPI-mediated efferocytosis provoked anti-inflammatory responses. Taken together, our work suggests that co-receptors are dispensable for tethering receptor-induced efferocytosis and that tethering receptors mediate the engulfment of apoptotic cells through multiple engulfment signaling pathways. The removal of apoptotic cells, known as efferocytosis, is a series of arranged events from the recruitment of phagocytes to sites where apoptotic cells are generated to the digestion of apoptotic cells by phagocytes.1, 2, 3 One of the key steps during efferocytosis is the recognition of dying cells by phagocytes. Phagocytes can detect apoptotic cells by the direct or indirect association of multiple receptors on phagocytes with ligands on apoptotic cells.4, 5, 6, 7, 8, 9 Some receptors on the surface of phagocytic cells not only bind to apoptotic cells but also transduce apoptotic cell recognition signals into phagocytes in order to mediate the ingestion of apoptotic cells. For instance, brain-specific angiogenesis inhibitor 1 (BAI1) and stabilin-2, which are phosphatidylserine (PtdSer) receptors, recognize PtdSer on apoptotic cells and relay signals to the Elmo-Dock-Rac module and Gulp, respectively, via their cytoplasmic tails.8, 10, 11 By contrast, it has been suggested that other receptors, called tethering receptors, merely tether apoptotic cells to phagocytes without AC710 mediating downstream signal transduction, following which the internalization of apoptotic cells is mediated by the association of these receptors with co-receptors or other engulfment receptors located nearby.12, 13, 14, 15, 16 However, it is unclear whether co-receptors for tethering receptors exist in tethering receptor-mediated phagocytosis of apoptotic cells, and, if indeed they do, if they are indispensable because of this procedure. One intriguing quality of tethering receptors can be they have cytoplasmic tails missing any signaling motifs or are anchored via glycophosphatidylinositol (GPI) towards the external leaflet from the plasma membrane. For instance, Tim-4, a PtdSer receptor with a brief cytoplasmic tail that promotes the engulfment of apoptotic cells from the binding of its IgV site to PtdSer on apoptotic cells, does not have signaling motifs in its cytoplasmic tail. It’s been known that neither the cytoplasmic tail nor the transmembrane area of Tim-4 is vital for Tim-4-mediated engulfment of apoptotic cells. Appropriately, it functions like a tethering receptor to protected apoptotic cells on phagocytes.9, 14 Compact disc14 is situated in the exofacial leaflet from the plasma membrane through its GPI anchor, which rules out the chance that it mediates direct signal transduction into phagocytes after binding to apoptotic cells. As a result, it is regarded as a tethering receptor also.15 Phospholipids such as for example PtdSer and phosphatidylcholine (PtdCho) are unequally distributed between your inner and outer leaflets from the plasma membrane in the standard state. For example, uncharged phospholipids such as for example PtdCho and sphingomyelin can be found in the outer leaflet mainly, whereas favorably or negatively charged phospholipids (such as phosphatidylethanolamine or PtdSer, respectively) are restricted to the inner leaflet facing the cytosol.17, 18, 19 However, this asymmetric distribution of phospholipids in the plasma membrane is disrupted during apoptosis. In the plasma membrane of apoptotic cells, PtdSer is exposed to the outer leaflet of the plasma membrane by the activity of scramblases and flippases.18, AC710 20, 21 Thus, exposed PtdSer is a hallmark of apoptotic cells and is the best characterized Esam ligand on apoptotic cells for efferocytosis. PtdSer on the surface of apoptotic cells can be recognized by various PtdSer-sensing membrane proteins on phagocytes, collectively called PtdSer receptors, including tethering receptors. Besides PtdSer receptors, many PtdSer-binding proteins have been identified. These proteins are involved in various biological processes AC710 such as blood coagulation, synaptic vesicle fusion, membrane scaffolding, and signal transduction.22 One of the best known proteins is annexin A5, which has been extensively studied as a PtdSer-binding protein. Annexin A5 belongs to the family of annexins, which are characterized by.