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D Monard - One of the best experts on this subject based on the ideXlab platform.
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the phosphatidylethanolamine binding protein is the prototype of a novel family of serine protease inhibitors
Journal of Biological Chemistry, 2001Co-Authors: Ulrich Hengst, Hugo Albrecht, Daniel Hess, D MonardAbstract:Abstract Serine proteases are involved in many processes in the nervous system and specific inhibitors tightly control their proteolytic activity. Thrombin is thought to play a role in tissue development and homeostasis. To date, protease Nexin-1 is the only known endogenous protease inhibitor that specifically interferes with thrombotic activity and is expressed in the brain. In this study, we report the detection of a novel thrombin inhibitory activity in the brain of protease Nexin-1(− /− ) mice. Purification and subsequent analysis by tandem mass spectrometry identified this protein as the phosphatidylethanolamine-binding protein (PEBP). We demonstrate that PEBP exerts inhibitory activity against several serine proteases including thrombin, neuropsin, and chymotrypsin, whereas trypsin, tissue type plasminogen activator, and elastase are not affected. Since PEBP does not share significant homology with other serine protease inhibitors, our results define it as the prototype of a novel class of serine protease inhibitors. PEBP immunoreactivity is found on the surface of Rat-1 fibroblast cells and although its sequence contains no secretion signal, PEBP-H6can be purified from the conditioned medium upon recombinant expression.
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glia derived Nexin protease Nexin 1 is expressed by a subset of neurons in the rat brain
Journal of Neuroscience Research, 1994Co-Authors: E Reinhard, H S Suidan, A Pavlik, D MonardAbstract:Glia-derived Nexin/protease Nexin-1 (GDN/PN-1) is a serine protease inhibitor that is secreted by glial cells and fibroblasts in culture. In the adult mammalian nervous system it has been shown to be expressed in the olfactory system and by some glial cells in response to neuronal injury. In situ hybridization and immunocytochemical studies were performed to identify the structures expressing GDN/PN-1 in the developing and adult rat brain. In contrast to a transient widespread expression during pre- and postnatal development, some brain structures constitutively express GDN/PN-1. These include the olfactory nerve layer of the olfactory bulb, basal forebrain, striatum, pyramidal neurons of layer V in the cortex, thalamic nuclei, pars compacta of the substantia nigra, inferior and superior colliculi, and deep cerebellar nuclei. All of these parts, excluding the olfactory nerve layer, are characterized by a high neuronal cell density. Neurons in these regions were immunoreactive for GDN/PN-1. Furthermore GDN/PN-1 expression in cell lines showed that the active protein was synthesized and secreted from B104 but not from NB2a neuroblastoma cells. Although GDN/PN-1 has only been reported to be synthesized by glia, the results presented here demonstrate that in addition, a subset of neurons express this protease inhibitor.
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the prolonged presence of glia derived Nexin an endogenous protease inhibitor in the hippocampus after ischemia induced delayed neuronal death
Neuroscience, 1992Co-Authors: M C Hoffmann, E Reinhard, Cordula Nitsch, A L Scotti, D MonardAbstract:The presence of glia-derived Nexin and glia fibrillary acidic protein (GFAP) was investigated in the hippocampus of Mongolian gerbils (Meriones unguiculatus) after transient forebrain ischemia. Bilateral clamping of the common carotid arteries for 7 min resulted in selective degeneration of CA1 pyramidal cells after a delay of three to four days, the so-called delayed neuronal death. Immunoreactivity for glia-derived Nexin was found in astrocytes of all CA1 layers and was detectable until day 90 (the longest survival time studied). Astroglial reactivity was demonstrated in parallel by staining for GFAP. The co-localization of glia-derived Nexin and GFAP was confirmed by double immunocytochemistry. Ultrastructural studies showed the exclusive presence of glia-derived Nexin in astrocytes, in the vicinity of degenerating and preserved neuronal structures. Perivascular glia was intensely stained, but endothelial cells were devoid of immunoreactivity. Glia-derived Nexin is a potent protease inhibitor with in vitro neurite-promoting activity. During adulthood, it is mainly present in the olfactory system, where receptor neurons are constantly being replaced. The ability of astrocytes to renew the expression of glia-derived Nexin after selective delayed neuronal death and the prolonged presence of the protease inhibitor in a zone where degeneration occurs in the immediate neighborhood of preserved neuronal elements indicate that glia-derived Nexin may play a role in structural rearrangements of the central nervous system.
Peter J. Cullen - One of the best experts on this subject based on the ideXlab platform.
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sorting Nexin 21 is a scaffold for the endosomal recruitment of huntingtin
Journal of Cell Science, 2018Co-Authors: Chris M Danson, Neil David Pearson, Kate J Heesom, Peter J. CullenAbstract:ABSTRACT The endo-lysosomal network serves an essential role in determining the fate of endocytosed transmembrane proteins and their associated proteins and lipids. Sorting Nexins (SNXs) play a central role in the functional organisation of this network. Comprising over 30 proteins in humans, SNXs are classified into sub-groups based on the presence of additional functional domains. Sorting Nexin-20 (SNX20) and sorting Nexin-21 (SNX21) comprise the SNX-PXB proteins. The presence of a predicted protein-protein interaction domain, termed the PX-associated B (PXB) domain, has led to the proposal that they function as endosome-associated scaffolds. Here, we used unbiased quantitative proteomics to define the SNX21 interactome. We reveal that the N-terminal extension of SNX21 interacts with huntingtin (Htt) whereas the PXB domain appears to associate with septins, a family of cytoskeletal- and membrane-associated proteins. In establishing that these interactions are sufficient for SNX21 to recruit Htt and septins on to an endosomal population, we reveal a scaffolding function for this sorting Nexin. Our work paves the way for a more-detailed mechanistic analysis of the role(s) of the SNX-PXB proteins in endosomal biology.
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SNX4 coordinates endosomal sorting of TfnR with dynein-mediated transport into the endocytic recycling compartment
Nature Cell Biology, 2007Co-Authors: Colin J Traer, Jez G. Carlton, Thomas Wassmer, Naomi Attar, Jacqueline Oakley, Anna C. Rutherford, Krysten J. Palmer, Joachim Kremerskothen, David J. Stephens, Peter J. CullenAbstract:SNX-BAR proteins are a sub-family of sorting Nexins implicated in endosomal sorting. Here, we establish that through its phox homology (PX) and Bin–Amphiphysin–Rvs (BAR) domains, sorting Nexin-4 (SNX4) is associated with tubular and vesicular elements of a compartment that overlaps with peripheral early endosomes and the juxtanuclear endocytic recycling compartment (ERC). Suppression of SNX4 perturbs transport between these compartments and causes lysosomal degradation of the transferrin receptor (TfnR). Through an interaction with KIBRA, a protein previously shown to bind dynein light chain 1, we establish that SNX4 associates with the minus end-directed microtubule motor dynein. Although suppression of KIBRA and dynein perturbs early endosome-to-ERC transport, TfnR sorting is maintained. We propose that by driving membrane tubulation, SNX4 coordinates iterative, geometric-based sorting of the TfnR with the long-range transport of carriers from early endosomes to the ERC. Finally, these data suggest that by associating with molecular motors, SNX-BAR proteins may coordinate sorting with carrier transport between donor and recipient membranes.
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a loss of function screen reveals snx5 and snx6 as potential components of the mammalian retromer
Journal of Cell Science, 2006Co-Authors: Thomas Wassmer, Miriam V Bujny, Naomi Attar, Colin J Traer, Jacqueline Oakley, Peter J. CullenAbstract:The mammalian retromer is a multimeric protein complex involved in mediating endosome-to-trans-Golgi-network retrograde transport of the cation-independent mannose-6-phosphate receptor. The retromer is composed of two subcomplexes, one containing SNX1 and forming a membrane-bound coat, the other comprising VPS26, VPS29 and VPS35 and being cargo-selective. In yeast, an additional sorting Nexin--Vps17p--is a component of the membrane bound coat. It remains unclear whether the mammalian retromer requires a functional equivalent of Vps17p. Here, we have used an RNAi loss-of-function screen to examine whether any of the other 30 mammalian sorting Nexins are required for retromer-mediated endosome-to-trans-Golgi-network retrieval of the cation-independent mannose-6-phosphate receptor. Using this screen, we identified two proteins, SNX5 and SNX6, that, when suppressed, induced a phenotype similar to that observed upon suppression of known retromer components. Whereas SNX5 and SNX6 colocalised with SNX1 on early endosomes, in immunoprecipitation experiments only SNX6 appeared to exist in a complex with SNX1. Interestingly, suppression of SNX5 and/or SNX6 resulted in a significant loss of SNX1, an effect that seemed to result from post-translational regulation of the SNX1 level. Such data suggest that SNX1 and SNX6 exist in a stable, endosomally associated complex that is required for retromer-mediated retrieval of the cation-independent mannose-6-phosphate receptor. SNX5 and SNX6 may therefore constitute functional equivalents of Vps17p in mammals.
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sorting Nexin 1 mediates tubular endosome to tgn transport through coincidence sensing of high curvature membranes and 3 phosphoinositides
Current Biology, 2004Co-Authors: Jez G. Carlton, Harry Mellor, Miriam V Bujny, Brian J Peter, Viola Oorschot, Anna Rutherford, Judith Klumperman, Harvey T Mcmahon, Peter J. CullenAbstract:Abstract Background: Sorting Nexins (SNXs) are phox homology (PX) domain-containing proteins thought to regulate endosomal sorting of internalized receptors. The prototypical SNX is sorting Nexin-1 (SNX1), a protein that through its PX domain binds phosphatidylinositol 3-monophosphate [PtdIns(3)P] and phosphatidylinositol 3,5-bisphosphate [PtdIns(3,5)P 2 ]. SNX1 is associated with early endosomes, from where it has been proposed to regulate the degradation of internalized epidermal growth factor (EGF) receptors through modulating endosomal-to-lysosomal sorting. Results: We show here that SNX1 contains a BAR (Bin/Amphiphysin/Rvs) domain, a membrane binding domain that endows SNX1 with the ability to form dimers and to sense membrane curvature. We present evidence that through coincidence detection, the BAR and PX domains efficiently target SNX1 to a microdomain of the early endosome defined by high curvature and the presence of 3-phosphoinositides. In addition, we show that the BAR domain endows SNX1 with an ability to tubulate membranes in-vitro and drive the tubulation of the endosomal compartment in-vivo. Using RNA interference (RNAi), we establish that SNX1 does not play a role in EGF or transferrin receptor sorting; rather it specifically perturbs endosome-to- trans Golgi network (TGN) transport of the cation-independent mannose-6-phosphate receptor (CI-MPR). Our data support an evolutionarily conserved function for SNX1 from yeast to mammals and provide functional insight into the molecular mechanisms underlying lipid-mediated protein targeting and tubular-based protein sorting. Conclusions: We conclude that through coincidence detection SNX1 associates with a microdomain of the early endosome—characterized by high membrane curvature and the presence of 3-phosphoinositides—from where it regulates tubular-based endosome-to-TGN retrieval of the CI-MPR.
Fuhai Ji - One of the best experts on this subject based on the ideXlab platform.
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reliability of a new 4th generation flotrac algorithm to track cardiac output changes in patients receiving phenylephrine
Journal of Clinical Monitoring and Computing, 2015Co-Authors: Neal Fleming, Fuhai Ji, Jian Li, David D RoseAbstract:Phenylephrine is often used to treat intra-operative hypotension. Previous studies have shown that the FloTrac cardiac monitor may overestimate cardiac output (CO) changes following phenylephrine administration. A new algorithm (4th generation) has been developed to improve performance in this setting. We performed a prospective observational study to assess the effects of phenylephrine administration on CO values measured by the 3rd and 4th generation FloTrac algorithms. 54 patients were enrolled in this study. We used the Nexfin, a pulse contour method shown to be insensitive to vasopressor administration, as the reference method. Radial arterial pressures were recorded continuously in patients undergoing surgery. Phenylephrine administration times were documented. Arterial pressure recordings were subsequently analyzed offline using three different pulse contour analysis algorithms: FloTrac 3rd generation (G3), FloTrac 4th generation (G4), and Nexfin (nf). One minute of hemodynamic measurements was analyzed immediately before phenylephrine administration and then repeated when the mean arterial pressure peaked. A total of 157 (4.6 ± 3.2 per patient, range 1–15) paired sets of hemodynamic recordings were analyzed. Phenylephrine induced a significant increase in stroke volume (SV) and CO with the FloTrac G3, but not with FloTrac G4 or Nexfin algorithms. Agreement between FloTrac G3 and Nexfin was: 0.23 ± 1.19 l/min and concordance was 51.1 %. In contrast, agreement between FloTrac G4 and Nexfin was: 0.19 ± 0.86 l/min and concordance was 87.2 %. In conclusion, the pulse contour method of measuring CO, as implemented in FloTrac 4th generation algorithm, has significantly improved its ability to track the changes in CO induced by phenylephrine.
Michael Overduin - One of the best experts on this subject based on the ideXlab platform.
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Phosphorylation of conserved phosphoinositide binding pocket regulates sorting Nexin membrane targeting.
Nature communications, 2018Co-Authors: Marc Lenoir, Jean Gruenberg, Cansel Ustunel, Sandya Rajesh, Jaswant Kaur, Dimitri Moreau, Michael OverduinAbstract:Sorting Nexins anchor trafficking machines to membranes by binding phospholipids. The paradigm of the superfamily is sorting Nexin 3 (SNX3), which localizes to early endosomes by recognizing phosphatidylinositol 3-phosphate (PI3P) to initiate retromer-mediated segregation of cargoes to the trans-Golgi network (TGN). Here we report the solution structure of full length human SNX3, and show that PI3P recognition is accompanied by bilayer insertion of a proximal loop in its extended Phox homology (PX) domain. Phosphoinositide (PIP) binding is completely blocked by cancer-linked phosphorylation of a conserved serine beside the stereospecific PI3P pocket. This "PIP-stop" releases endosomal SNX3 to the cytosol, and reveals how protein kinases control membrane assemblies. It constitutes a widespread regulatory element found across the PX superfamily and throughout evolution including of fungi and plants. This illuminates the mechanism of a biological switch whereby structured PIP sites are phosphorylated to liberate protein machines from organelle surfaces.
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Phosphorylation of conserved phosphoinositide binding pocket regulates sorting Nexin membrane targeting
Nature Publishing Group, 2018Co-Authors: Marc Lenoir, Jean Gruenberg, Cansel Ustunel, Sandya Rajesh, Jaswant Kaur, Dimitri Moreau, Michael OverduinAbstract:Sorting Nexin 3 (SNX3) is a phosphatidylinositol 3-phosphate binding protein that localizes to early endosomes. Here the authors use NMR to resolve SNX3′s membrane interactions, revealing that membrane binding is regulated through phosphorylation of a conserved serine by its lipid recognition site
David D Rose - One of the best experts on this subject based on the ideXlab platform.
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reliability of a new 4th generation flotrac algorithm to track cardiac output changes in patients receiving phenylephrine
Journal of Clinical Monitoring and Computing, 2015Co-Authors: Neal Fleming, Fuhai Ji, Jian Li, David D RoseAbstract:Phenylephrine is often used to treat intra-operative hypotension. Previous studies have shown that the FloTrac cardiac monitor may overestimate cardiac output (CO) changes following phenylephrine administration. A new algorithm (4th generation) has been developed to improve performance in this setting. We performed a prospective observational study to assess the effects of phenylephrine administration on CO values measured by the 3rd and 4th generation FloTrac algorithms. 54 patients were enrolled in this study. We used the Nexfin, a pulse contour method shown to be insensitive to vasopressor administration, as the reference method. Radial arterial pressures were recorded continuously in patients undergoing surgery. Phenylephrine administration times were documented. Arterial pressure recordings were subsequently analyzed offline using three different pulse contour analysis algorithms: FloTrac 3rd generation (G3), FloTrac 4th generation (G4), and Nexfin (nf). One minute of hemodynamic measurements was analyzed immediately before phenylephrine administration and then repeated when the mean arterial pressure peaked. A total of 157 (4.6 ± 3.2 per patient, range 1–15) paired sets of hemodynamic recordings were analyzed. Phenylephrine induced a significant increase in stroke volume (SV) and CO with the FloTrac G3, but not with FloTrac G4 or Nexfin algorithms. Agreement between FloTrac G3 and Nexfin was: 0.23 ± 1.19 l/min and concordance was 51.1 %. In contrast, agreement between FloTrac G4 and Nexfin was: 0.19 ± 0.86 l/min and concordance was 87.2 %. In conclusion, the pulse contour method of measuring CO, as implemented in FloTrac 4th generation algorithm, has significantly improved its ability to track the changes in CO induced by phenylephrine.
