The Experts below are selected from a list of 207 Experts worldwide ranked by ideXlab platform
Shigekazu Nagata - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of a human plasma membrane phospholipid flippase
Journal of Biological Chemistry, 2020Co-Authors: Hanayo Nakanishi, Katsumasa Irie, Katsumori Segawa, Kazuya Hasegawa, Yoshinori Fujiyoshi, Shigekazu Nagata, Kazuhiro AbeAbstract:ATP11C, a member of the P4-ATPase flippase, translocates Phosphatidylserine from the outer to the inner plasma membrane leaflet, and maintains the asymmetric distribution of Phosphatidylserine in the living cell. We present the crystal structures of a human plasma membrane flippase, ATP11C-CDC50A complex, in a stabilized E2P conformation. The structure revealed a deep longitudinal crevice along transmembrane helices continuing from the cell surface to the phospholipid occlusion site in the middle of the membrane. We observed that the extension of the crevice on the exoplasmic side is open, and the complex is therefore in an outward-open E2P state, similar to a recently reported cryo-EM structure of yeast flippase Drs2p-Cdc50p complex. We noted extra densities, most likely bound Phosphatidylserines, in the crevice and in its extension to the extracellular side. One was close to the Phosphatidylserine occlusion site as previously reported for the human ATP8A1-CDC50A complex, and the other in a cavity at the surface of the exoplasmic leaflet of the bilayer. Substitutions in either of the binding sites or along the path between them impaired specific ATPase and transport activities. These results provide evidence that the observed crevice is the conduit along which Phosphatidylserine traverses from the outer leaflet to its occlusion site in the membrane and suggest that the exoplasmic cavity is important for phospholipid recognition. They also yield insights into how Phosphatidylserine is incorporated from the outer leaflet of the plasma membrane into the transmembrane.
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crystal structure of a human plasma membrane phospholipid flippase
bioRxiv, 2019Co-Authors: Hanayo Nakanishi, Katsumasa Irie, Katsumori Segawa, Kazuya Hasegawa, Yoshinori Fujiyoshi, Shigekazu Nagata, Kazuhiro AbeAbstract:ATP11C, a member of P4-ATPase flippase, exclusively translocates Phosphatidylserine from the outer to the inner leaflets of the plasma membrane, and maintains the asymmetric distribution of Phosphatidylserine in the living cell. However, the mechanisms by which ATP11C translocates Phosphatidylserine remain elusive. Here we show the crystal structures of a human plasma membrane flippase, ATP11C-CDC50A complex, in an outward-open E2P conformation. Two Phosphatidylserine molecules are in a conduit that continues from the cell surface to the occlusion site in the middle of the membrane. Mutations in either of the phosphotidylserine binding sites or along the pathway between significantly impairs specific ATPase and transport activities. We propose a model for Phosphatidylserine translocation from the outer to the inner leaflet of the plasma membrane.
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Role of lactadherin in the clearance of Phosphatidylserine-expressing red blood cells.
Transfusion, 2008Co-Authors: Swapan K. Dasgupta, Shigekazu Nagata, Hanan Abdel-monem, Prasenjit Guchhait, Perumal ThiagarajanAbstract:BACKGROUND: In red blood cells (RBCs) anionic phospholipids, such as Phosphatidylserine, are present in the inner leaflet of the membrane bilayer. Exposure of Phosphatidylserine occurs during senescence and during long-term storage of RBCs and is considered as the tag for removal from the circulation by macrophages. Lactadherin is a Phosphatidylserine-binding glycoprotein secreted by macrophages that promotes the engulfment of Phosphatidylserine-expressing apoptotic lymphocytes. This study investigates the role of lactadherin in the phagocytosis of Phosphatidylserine-expressing RBCs. STUDY DESIGN AND METHODS: Transbilayer movement of Phosphatidylserine was induced in RBCs either by storage beyond 30 days or by treatment with calcium ionophore A23187 and N-ethylmaleimide. Phosphatidylserine-expressing RBCs were incubated with phorbol ester–stimulated THP-1, and phagocytosis was determined by measuring the pseudoperoxidase activity of hemoglobin. The in vivo clearance of Phosphatidylserine-enriched RBCs was measured in lactadherin-deficient mice and in their littermate controls. RESULTS: Lactadherin promoted phagocytosis of Phosphatidylserine-expressing RBCs by macrophages in a concentration-dependent manner. Splenic macrophages from lactadherin-deficient mice had diminished capacity to phagocytose Phosphatidylserine-expressing RBCs. The life span of RBCs in lactadherin-deficient mice was similar to wild-type littermate controls in vivo. However, when an excess of Phosphatidylserine-expressing RBCs were infused, there was only a mild impairment in the clearance in lactadherin-deficient mice compared to wild-type littermate controls. CONCLUSION: These results show that clearance of Phosphatidylserine-expressing RBCs is not diminished in a significant way in lactadherin-deficient mice under physiologic conditions and suggest the presence of redundant pathways.
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Identification of Tim4 as a Phosphatidylserine receptor
Nature, 2007Co-Authors: Masanori Miyanishi, Kazutoshi Tada, Masato Koike, Yasuo Uchiyama, Toshio Kitamura, Shigekazu NagataAbstract:During programmed cell death in multicellular organisms, a large number of cells are engulfed by macrophages, thus avoiding the release of noxious materials from the dying cells. These 'apoptotic' cells expose Phosphatidylserine (PS) on their surface as an 'eat-me' signal. Miyanishi et al. show that the receptors Tim4 and Tim1 are implicated in phagocyte recognition of PS, while Park et al. show that the BAI1 protein is a receptor for PS in mammalian macrophages. Apoptotic cells expose Phosphatidylserine as an 'eat-me' signal for macrophages. This paper shows that the receptors Tim4 and Tim1 are implicated in phagocyte recognition of Phosphatidylserine. In programmed cell death, a large number of cells undergo apoptosis, and are engulfed by macrophages to avoid the release of noxious materials from the dying cells1,2. In definitive erythropoiesis, nuclei are expelled from erythroid precursor cells and are engulfed by macrophages. Phosphatidylserine is exposed on the surface of apoptotic cells3 and on the nuclei expelled from erythroid precursor cells4; it works as an ‘eat me’ signal for phagocytes5,6. Phosphatidylserine is also expressed on the surface of exosomes involved in intercellular signalling7. Here we established a library of hamster monoclonal antibodies against mouse peritoneal macrophages, and found an antibody that strongly inhibited the Phosphatidylserine-dependent engulfment of apoptotic cells. The antigen recognized by the antibody was identified by expression cloning as a type I transmembrane protein called Tim4 (T-cell immunoglobulin- and mucin-domain-containing molecule; also known as Timd4)8. Tim4 was expressed in Mac1+ cells in various mouse tissues, including spleen, lymph nodes and fetal liver. Tim4 bound apoptotic cells by recognizing Phosphatidylserine via its immunoglobulin domain. The expression of Tim4 in fibroblasts enhanced their ability to engulf apoptotic cells. When the anti-Tim4 monoclonal antibody was administered into mice, the engulfment of apoptotic cells by thymic macrophages was significantly blocked, and the mice developed autoantibodies. Among the other Tim family members, Tim1, but neither Tim2 nor Tim3, specifically bound Phosphatidylserine. Tim1- or Tim4-expressing Ba/F3 B cells were bound by exosomes via Phosphatidylserine, and exosomes stimulated the interaction between Tim1 and Tim4. These results indicate that Tim4 and Tim1 are Phosphatidylserine receptors for the engulfment of apoptotic cells, and may also be involved in intercellular signalling in which exosomes are involved.
Perumal Thiagarajan - One of the best experts on this subject based on the ideXlab platform.
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Platelet senescence and Phosphatidylserine exposure
Transfusion, 2010Co-Authors: Swapan K. Dasgupta, Hanan Abdel-monem, Anthony Prakasam, Eduardo Rios Argaiz, Jose Emmanel Chedid Mercado, Hector Omar Elizondo Maul, Jorge Garza, Ana Bety Enriquez, Michael Andreeff, Perumal ThiagarajanAbstract:Background The exposure of Phosphatidylserine occurs during platelet activation and during in vitro storage. Phosphatidylserine exposure also occurs during apoptosis following the release of mitochondrial cytochrome c. We have examined the role of cytochrome c release, mitochondrial membrane potential (ΔΨm), and cyclophilin D (CypD) in Phosphatidylserine exposure due to activation and storage.
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Platelet Senescence and Phosphatidylserine Exposure.
Blood, 2009Co-Authors: Swapan K. Dasgupta, Hanan Abdel-monem, Anthony Prakasam, Perumal ThiagarajanAbstract:Abstract 2117 Poster Board II-94 The transbilayer movement of Phosphatidylserine from the inner to the outer leaflet of membrane bilayer occurs during platelet activation and is essential for platelet procoagulant activity. Expression of Phosphatidylserine also occurs in platelets stored more than 5 days in the blood bank. We have examined the role of mitochondrial pathways in activation-induced platelet procoagulant activity and during in vitro senescence. Collagen and thrombin-induced exposure of Phosphatidylserine is associated with a decrease in mitochondrial membrane potential (MMP). Cyclosporin A, a known inhibitor of cyclophilin D, inhibited Phosphatidylserine exposure and the loss of MMP. Consistent with this, platelets from CypD deficient mice had decreased loss of MMP and impaired Phosphatidylserine exposure when treated with a combination of thrombin and collagen. However, there is no release of cytochrome c into the cytosol. Also there is no activation of caspase-3 or Rho associated kinase I (ROCK1). In sharp contrast to this, in platelets stored for more than 5 days, Phosphatidylserine exposure is found to be associated with caspase-3 and ROCK1 activation. To delineate the mechanism we used ABT737, a BH3 mimetic that induces mitochondrial pathway of apoptosis via Bcl-2 family of proteins. ABT737-induced Phosphatidylserine exposure was also associated with cytochrome c release, caspase-3 and ROCKI activation and it is unaffected in platelets isolated from CypD knock out mice or in platelets treated with cyclosporine A prior to activation. Caspase-3 inhibitor (z-DVED-fmk and ROCK1 inhibitor Y-27632 reduced ABT-737 induced Phosphatidylserine expression. Our results also show that activation-induced Phosphatidylserine exposure is dependent on mitochondrial membrane depolarization and CypD but is independent of cytochrome c release or caspase-3 or ROCK1 activation. In contrast, the Phosphatidylserine exposure in stored platelets occurs via mitochondrial cytochrome c release, caspase-3 activation and ROCKI activation and does not depend on mitochondrial membrane depolarization or CypD. Disclosures: No relevant conflicts of interest to declare.
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Role of lactadherin in the clearance of Phosphatidylserine-expressing red blood cells.
Transfusion, 2008Co-Authors: Swapan K. Dasgupta, Shigekazu Nagata, Hanan Abdel-monem, Prasenjit Guchhait, Perumal ThiagarajanAbstract:BACKGROUND: In red blood cells (RBCs) anionic phospholipids, such as Phosphatidylserine, are present in the inner leaflet of the membrane bilayer. Exposure of Phosphatidylserine occurs during senescence and during long-term storage of RBCs and is considered as the tag for removal from the circulation by macrophages. Lactadherin is a Phosphatidylserine-binding glycoprotein secreted by macrophages that promotes the engulfment of Phosphatidylserine-expressing apoptotic lymphocytes. This study investigates the role of lactadherin in the phagocytosis of Phosphatidylserine-expressing RBCs. STUDY DESIGN AND METHODS: Transbilayer movement of Phosphatidylserine was induced in RBCs either by storage beyond 30 days or by treatment with calcium ionophore A23187 and N-ethylmaleimide. Phosphatidylserine-expressing RBCs were incubated with phorbol ester–stimulated THP-1, and phagocytosis was determined by measuring the pseudoperoxidase activity of hemoglobin. The in vivo clearance of Phosphatidylserine-enriched RBCs was measured in lactadherin-deficient mice and in their littermate controls. RESULTS: Lactadherin promoted phagocytosis of Phosphatidylserine-expressing RBCs by macrophages in a concentration-dependent manner. Splenic macrophages from lactadherin-deficient mice had diminished capacity to phagocytose Phosphatidylserine-expressing RBCs. The life span of RBCs in lactadherin-deficient mice was similar to wild-type littermate controls in vivo. However, when an excess of Phosphatidylserine-expressing RBCs were infused, there was only a mild impairment in the clearance in lactadherin-deficient mice compared to wild-type littermate controls. CONCLUSION: These results show that clearance of Phosphatidylserine-expressing RBCs is not diminished in a significant way in lactadherin-deficient mice under physiologic conditions and suggest the presence of redundant pathways.
In-san Kim - One of the best experts on this subject based on the ideXlab platform.
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Stabilin Receptors: Role as Phosphatidylserine Receptors
Biomolecules, 2019Co-Authors: Seung Yoon Park, In-san KimAbstract:Phosphatidylserine is a membrane phospholipid that is localized to the inner leaflet of the plasma membrane. Phosphatidylserine externalization to the outer leaflet of the plasma membrane is an important signal for various physiological processes, including apoptosis, platelet activation, cell fusion, lymphocyte activation, and regenerative axonal fusion. Stabilin-1 and stabilin-2 are membrane receptors that recognize Phosphatidylserine on the cell surface. Here, we discuss the functions of Stabilin-1 and stabilin-2 as Phosphatidylserine receptors in apoptotic cell clearance (efferocytosis) and cell fusion, and their ligand-recognition and signaling pathways.
Swapan K. Dasgupta - One of the best experts on this subject based on the ideXlab platform.
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Platelet senescence and Phosphatidylserine exposure
Transfusion, 2010Co-Authors: Swapan K. Dasgupta, Hanan Abdel-monem, Anthony Prakasam, Eduardo Rios Argaiz, Jose Emmanel Chedid Mercado, Hector Omar Elizondo Maul, Jorge Garza, Ana Bety Enriquez, Michael Andreeff, Perumal ThiagarajanAbstract:Background The exposure of Phosphatidylserine occurs during platelet activation and during in vitro storage. Phosphatidylserine exposure also occurs during apoptosis following the release of mitochondrial cytochrome c. We have examined the role of cytochrome c release, mitochondrial membrane potential (ΔΨm), and cyclophilin D (CypD) in Phosphatidylserine exposure due to activation and storage.
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Platelet Senescence and Phosphatidylserine Exposure.
Blood, 2009Co-Authors: Swapan K. Dasgupta, Hanan Abdel-monem, Anthony Prakasam, Perumal ThiagarajanAbstract:Abstract 2117 Poster Board II-94 The transbilayer movement of Phosphatidylserine from the inner to the outer leaflet of membrane bilayer occurs during platelet activation and is essential for platelet procoagulant activity. Expression of Phosphatidylserine also occurs in platelets stored more than 5 days in the blood bank. We have examined the role of mitochondrial pathways in activation-induced platelet procoagulant activity and during in vitro senescence. Collagen and thrombin-induced exposure of Phosphatidylserine is associated with a decrease in mitochondrial membrane potential (MMP). Cyclosporin A, a known inhibitor of cyclophilin D, inhibited Phosphatidylserine exposure and the loss of MMP. Consistent with this, platelets from CypD deficient mice had decreased loss of MMP and impaired Phosphatidylserine exposure when treated with a combination of thrombin and collagen. However, there is no release of cytochrome c into the cytosol. Also there is no activation of caspase-3 or Rho associated kinase I (ROCK1). In sharp contrast to this, in platelets stored for more than 5 days, Phosphatidylserine exposure is found to be associated with caspase-3 and ROCK1 activation. To delineate the mechanism we used ABT737, a BH3 mimetic that induces mitochondrial pathway of apoptosis via Bcl-2 family of proteins. ABT737-induced Phosphatidylserine exposure was also associated with cytochrome c release, caspase-3 and ROCKI activation and it is unaffected in platelets isolated from CypD knock out mice or in platelets treated with cyclosporine A prior to activation. Caspase-3 inhibitor (z-DVED-fmk and ROCK1 inhibitor Y-27632 reduced ABT-737 induced Phosphatidylserine expression. Our results also show that activation-induced Phosphatidylserine exposure is dependent on mitochondrial membrane depolarization and CypD but is independent of cytochrome c release or caspase-3 or ROCK1 activation. In contrast, the Phosphatidylserine exposure in stored platelets occurs via mitochondrial cytochrome c release, caspase-3 activation and ROCKI activation and does not depend on mitochondrial membrane depolarization or CypD. Disclosures: No relevant conflicts of interest to declare.
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Role of lactadherin in the clearance of Phosphatidylserine-expressing red blood cells.
Transfusion, 2008Co-Authors: Swapan K. Dasgupta, Shigekazu Nagata, Hanan Abdel-monem, Prasenjit Guchhait, Perumal ThiagarajanAbstract:BACKGROUND: In red blood cells (RBCs) anionic phospholipids, such as Phosphatidylserine, are present in the inner leaflet of the membrane bilayer. Exposure of Phosphatidylserine occurs during senescence and during long-term storage of RBCs and is considered as the tag for removal from the circulation by macrophages. Lactadherin is a Phosphatidylserine-binding glycoprotein secreted by macrophages that promotes the engulfment of Phosphatidylserine-expressing apoptotic lymphocytes. This study investigates the role of lactadherin in the phagocytosis of Phosphatidylserine-expressing RBCs. STUDY DESIGN AND METHODS: Transbilayer movement of Phosphatidylserine was induced in RBCs either by storage beyond 30 days or by treatment with calcium ionophore A23187 and N-ethylmaleimide. Phosphatidylserine-expressing RBCs were incubated with phorbol ester–stimulated THP-1, and phagocytosis was determined by measuring the pseudoperoxidase activity of hemoglobin. The in vivo clearance of Phosphatidylserine-enriched RBCs was measured in lactadherin-deficient mice and in their littermate controls. RESULTS: Lactadherin promoted phagocytosis of Phosphatidylserine-expressing RBCs by macrophages in a concentration-dependent manner. Splenic macrophages from lactadherin-deficient mice had diminished capacity to phagocytose Phosphatidylserine-expressing RBCs. The life span of RBCs in lactadherin-deficient mice was similar to wild-type littermate controls in vivo. However, when an excess of Phosphatidylserine-expressing RBCs were infused, there was only a mild impairment in the clearance in lactadherin-deficient mice compared to wild-type littermate controls. CONCLUSION: These results show that clearance of Phosphatidylserine-expressing RBCs is not diminished in a significant way in lactadherin-deficient mice under physiologic conditions and suggest the presence of redundant pathways.
Peter M. Henson - One of the best experts on this subject based on the ideXlab platform.
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cell corpse engulfment mediated by c elegans Phosphatidylserine receptor through ced 5 and ced 12
Science, 2003Co-Authors: Xiaochen Wang, Valerie A. Fadok, Ming Chia Lee, Keiko Gengyoando, Lichun Cheng, Duncan Ledwich, Pei Ken Hsu, Jiayun Chen, Bin Kuan Chou, Peter M. HensonAbstract:During apoptosis, Phosphatidylserine, which is normally restricted to the inner leaflet of the plasma membrane, is exposed on the surface of apoptotic cells and has been suggested to act as an “eat-me” signal to trigger phagocytosis. It is unclear how phagocytes recognize Phosphatidylserine. Recently, a putative Phosphatidylserine receptor (PSR) was identified and proposed to mediate recognition of Phosphatidylserine and phagocytosis. We report that psr-1, the Caenorhabditis elegans homolog of PSR, is important for cell corpse engulfment. In vitro PSR-1 binds preferentially Phosphatidylserine or cells with exposed Phosphatidylserine. In C. elegans, PSR-1 acts in the same cell corpse engulfment pathway mediated by intracellular signaling molecules CED-2 (homologous to the human CrkII protein), CED-5 (DOCK180), CED-10 (Rac GTPase), and CED-12 (ELMO), possibly through direct interaction with CED-5 and CED-12. Our findings suggest that PSR-1 is likely an upstream receptor for the signaling pathway containing CED-2, CED-5, CED-10, and CED-12 proteins and plays an important role in recognizing Phosphatidylserine during phagocytosis.
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A receptor for Phosphatidylserine-specific clearance of apoptotic cells
Nature, 2000Co-Authors: Valerie A. Fadok, Donna L. Bratton, R. Alan B. Ezekewitz, Alan Pearson, David M. Rose, Peter M. HensonAbstract:The culmination of apoptosis in vivo is phagocytosis of cellular corpses. During apoptosis, the asymmetry of plasma membrane phospholipids is lost, which exposes Phosphatidylserine externally. The phagocytosis of apoptotic cells can be inhibited stereospecifically by Phosphatidylserine and its structural analogues, but not by other anionic phospholipids, suggesting that Phosphatidylserine is specifically recognized. Using phage display, we have cloned a gene that appears to recognize Phosphatidylserine on apoptotic cells. Here we show that this gene, when transfected into B and T lymphocytes, enables them to recognize and engulf apoptotic cells in a Phosphatidylserine-specific manner. Flow cytometric analysis using a monoclonal antibody suggested that the protein is expressed on the surface of macrophages, fibroblasts and epithelial cells; this antibody, like Phosphatidylserine liposomes, inhibited the phagocytosis of apoptotic cells and, in macrophages, induced an anti-inflammatory state. This candidate Phosphatidylserine receptor is highly homologous to genes of unknown function in Caenorhabditis elegans and Drosophila melanogaster, suggesting that Phosphatidylserine recognition on apoptotic cells during their removal by phagocytes is highly conserved throughout phylogeny.