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BBA - Molecular Cell Research (v.1763, #11)
Special issue: The 9th European symposium on calcium-binding proteins in normal and transformed cells
by Jacques Haiech; Claus W. Heizmann; Joachim Krebs (pp. 1135-1135).
The evolution of calcium biochemistry by Robert J.P. Williams (pp. 1139-1146).
The role of calcium in evolution is best understood from a perspective based on its intrinsic value as a divalent cation able to bind and precipitate inorganic and organic anions rapidly. This binding can be useful or inhibitory. Now treatment of binding or precipitation has two different interests in biological cells. The first is thermodynamic, that is the stress is on systems biology and the second is structure, that is molecular biology. In evolution both have equal weight being connected through exchange. This paper outlines first the systems biology of the evolution of calcium functions from prokaryotes to animals with brains. The calcium ion was the only good available candidate in the environment for the functions it performs. The second section of the paper describes the evolution of the proteins which allow the messenger function. We have discussed elsewhere the structure/function relationships of the proteins. Overall the evolving and increasing involvement of calcium as possibly the major control messenger of events outside cells to action inside them is an inevitable feature of the nature of ecological, that is environmental/organism, evolution.
Keywords: Evolution; Calcium ion property; Calcium protein; Calcium cell concentration; Extracellular fluid; Calcium messenger
Emerging perspectives in store-operated Ca2+ entry: Roles of Orai, Stim and TRP by Jeremy T. Smyth; Wayne I. DeHaven; Bertina F. Jones; Jason C. Mercer; Mohamed Trebak; Guillermo Vazquez; James W. Putney Jr. (pp. 1147-1160).
Depletion of intracellular Ca2+ stores induces Ca2+ influx across the plasma membrane through store-operated channels (SOCs). This store-operated Ca2+ influx is important for the replenishment of the Ca2+ stores, and is also involved in many signaling processes by virtue of the ability of intracellular Ca2+ to act as a second messenger. For many years, the molecular identities of particular SOCs, as well as the signaling mechanisms by which these channels are activated, have been elusive. Recently, however, the mammalian proteins STIM1 and Orai1 were shown to be necessary for the activation of store-operated Ca2+ entry in a variety of mammalian cells. Here we present molecular, pharmacological, and electrophysiological properties of SOCs, with particular focus on the roles that STIM1 and Orai1 may play in the signaling processes that regulate various pathways of store-operated entry.
Keywords: Abbreviations; [Ca; 2+; ]; i; intracellular Ca; 2+; concentration; 2-APB; 2-aminoethoxydiphenylborane; AA; arachidonic acid; ARC; arachidonate-regulated channel; CCE; capacitative Ca; 2+; entry; CIF; Ca; 2; influx factor; DAG; diacylglycerol; ER; endoplasmic reticulum; EYFP; enhanced yellow fluorescent protein; I; CRAC; Ca; 2+; release-activated Ca; 2+; current; IP; 3; inositol 1,4,5-trisphosphate; IP; 3; R; inositol 1,4,5-trisphosphate receptor; OAG; 1-oleoyl-2-acetyl-; sn; -glycerol; PI; phosphoinositide; PIP; 2; phosphatidylinositol 4,5-bisphosphate; PKC; protein kinase C; PLA; 2; phospholipase A; 2; PLC; phospholipase C; ROC; receptor-operated channel; SAM; sterile-alpha motif; SCID; severe combined immunodeficiency; SERCA; sarco/endoplasmic reticulum Ca; 2+; -ATPase; SOC; store-operated channel; TRP; transient receptor potentialCalcium channel; TRP channel; Stim1; Orai1; Calcium-release-activated-calcium current; Store-operated channel; Capacitative calcium entry; Calcium oscillation
Calcium signals mediated by STIM and Orai proteins—A new paradigm in inter-organelle communication by Jonathan Soboloff; Maria A. Spassova; Marie A. Dziadek; Donald L. Gill (pp. 1161-1168).
In all cells Ca2+ signals are key to controlling a spectrum of cellular responses. Ca2+ signals activated by phospholipase C-coupled receptors have two components—rapid Ca2+ release from ER stores followed by slower Ca2+ entry from outside the cell. The coupling process between ER and PM to mediate this “store-operated� Ca2+ entry process has remained a molecular and mechanistic mystery. Through a combination of high throughput screening and molecular physiological approaches, the machinery and mechanism of this process have been elucidated. Two proteins are key to the coupling process. STIM1, a single spanning membrane protein with an unpaired Ca2+ binding EF-hand functions as the sensor of ER luminal Ca2+ and through redistribution in the ER transduces information directly to the PM. Orai1, a tetra-spanning PM protein, functions as the highly Ca2+ selective channel in the PM that is gated through interactions with the store-activated ER Ca2+ sensor. This molecular pas-de-deux between ER and PM components represents not only a crucial signaling pathway, but also a new paradigm in inter-organelle communication.
Keywords: Calcium; Calcium signals; Orai; STIM; Store-operated channels
Voltage-gated calcium channels in genetic diseases by Isabelle Bidaud; Alexandre Mezghrani; Leigh Anne Swayne; Arnaud Monteil; Philippe Lory (pp. 1169-1174).
Voltage-gated calcium channels (VGCCs) mediate calcium entry into excitable cells in response to membrane depolarization. During the past decade, our understanding of the gating and functions of VGCCs has been illuminated by the analysis of mutations linked to a heterogeneous group of genetic diseases called “calcium channelopathies�. Calcium channelopathies include muscular, neurological, cardiac and vision syndromes. Recent data suggest that calcium channelopathies result not only from electrophysiological defects but also from altered α1/CaV subunit protein processing, including folding, posttranslational modifications, quality control and trafficking abnormalities. Overall, functional analyses of VGCC mutations provide a more comprehensive view of the corresponding human disorders and offer important new insights into VGCC function. Ultimately, the understanding of these pathogenic channel mutations should lead to improved treatments of such hereditary diseases in humans.
Keywords: Calcium channelopathies; Hypokalemic periodic paralysis; Long QT syndrome; Ataxia; Migraine; Epilepsy; Autism
Exocytosis in neuroendocrine cells: New tasks for actin by Magali Malacombe; Marie-France Bader; Stéphane Gasman (pp. 1175-1183).
Most secretory cells undergoing calcium-regulated exocytosis in response to cell surface receptor stimulation display a dense subplasmalemmal actin network, which is remodeled during the exocytotic process. This review summarizes new insights into the role of the cortical actin cytoskeleton in exocytosis. Many earlier findings support the actin–physical-barrier model whereby transient depolymerization of cortical actin filaments permits vesicles to gain access to their appropriate docking and fusion sites at the plasma membrane. On the other hand, data from our laboratory and others now indicate that actin polymerization also plays a positive role in the exocytotic process. Here, we discuss the potential functions attributed to the actin cytoskeleton at each major step of the exocytotic process, including recruitment, docking and fusion of secretory granules with the plasma membrane. Moreover, we present actin-binding proteins, which are likely to link actin organization to calcium signals along the exocytotic pathway. The results cited in this review are derived primarily from investigations of the adrenal medullary chromaffin cell, a cell model that is since many years a source of information concerning the molecular machinery underlying exocytosis.
Keywords: Rho protein; Actin; Actin-binding protein; Exocytosis; Granule; Neuroendocrine cell
Calcium transients and calcium signalling during early neurogenesis in the amphibian embryo Xenopus laevis by Catherine Leclerc; Isabelle Néant; Sarah E. Webb; Andrew L. Miller; Marc Moreau (pp. 1184-1191).
Development of the vertebrate embryonic nervous system is characterized by a cascade of signalling events. In Xenopus, the initial step in this cascade results from signals emanating from the dorsal mesoderm that divert the fate of the ectoderm from an epidermal to a neural lineage. These signals include extracellular antagonists of the bone morphogenetic protein (BMP). Experiments performed with isolated ectoderm suggest that epidermis is induced by BMP, whereas neural fates arise by default following BMP inhibition; however, we show that this mechanism is not sufficient for neural determination. Ca2+ imaging of intact Xenopus embryos reveals patterns of Ca2+ transients in the dorsal ectoderm but not in the ventral ectoderm. These increases in intracellular calcium concentration ([Ca2+]i), which occur via the activation of dihydropyridine (DHP)-sensitive Ca2+ channels, are necessary and sufficient to orientate the ectodermal cells toward a neural fate. On the one hand, the treatments that antagonize the increase in [Ca2+]i, inhibit neuralization, while on the other hand, an artificial increase in [Ca2+]i, whatever its origin, neuralizes the ectoderm. Using these properties, we have constructed a subtractive cDNA library between untreated ectoderm and caffeine-treated ectoderm. The caffeine stimulates an increase in [Ca2+]i and thus orientates the cells towards the neural pathway. We have identified early Ca2+ target genes expressed in neural territories. One of these genes, an arginine methyl transferase, controls the expression of the early proneural gene , Zic3. Here, we discuss an alternative model where Ca2+ plays a central regulatory role in early neurogenesis. This model integrates the activation of a Ca2+-dependent signalling pathway due to an influx of Ca2+ through DHP-Ca2+ channels. While Ca2+ is required for neural determination, epidermal determination occurs when Ca2+-dependent signalling pathways are inactive.
Keywords: Calcium; DHP-channel; Neural determination; Xenopus laevis; Gene expression
Ca2+ signaling and early embryonic patterning during the Blastula and Gastrula Periods of Zebrafish and Xenopus development by Sarah E. Webb; Andrew L. Miller (pp. 1192-1208).
It has been proposed that Ca2+ signaling, in the form of pulses, waves and steady gradients, may play a crucial role in key pattern forming events during early vertebrate development [L.F. Jaffe, Organization of early development by calcium patterns, BioEssays 21 (1999) 657–667; M.J. Berridge, P. Lipp, M.D. Bootman, The versatility and universality of calcium signaling, Nat. Rev. Mol. Cell Biol. 1 (2000) 11–21; S.E. Webb, A.L. Miller, Calcium signalling during embryonic development, Nat. Rev. Mol. Cell Biol. 4 (2003) 539–551]. With reference to the embryos of zebrafish ( Danio rerio) and the frog, Xenopus laevis, we review the Ca2+ signals reported during the Blastula and Gastrula Periods. This developmental window encompasses the major pattern forming events of epiboly, involution, and convergent extension, which result in the establishment of the basic germ layers and body axes [C.B. Kimmel, W.W. Ballard, S.R. Kimmel, B. Ullmann, T.F. Schilling, Stages of embryonic development of the zebrafish, Dev. Dyn. 203 (1995) 253–310]. Data will be presented to support the suggestion that propagating waves (both long and short range) of Ca2+ release, followed by sequestration, may play a crucial role in: (1) Coordinating cell movements during these pattern forming events and (2) Contributing to the establishment of the basic embryonic axes, as well as (3) Helping to define the morphological boundaries of specific tissue domains and embryonic structures, including future organ anlagen [E. Gilland, A.L. Miller, E. Karplus, R. Baker, S.E. Webb, Imaging of multicellular large-scale rhythmic calcium waves during zebrafish gastrulation, Proc. Natl. Acad. Sci. USA 96 (1999) 157–161; J.B. Wallingford, A.J. Ewald, R.M. Harland, S.E. Fraser, Calcium signaling during convergent extension in Xenopus, Curr. Biol. 11 (2001) 652–661]. The various potential targets of these Ca2+ transients will also be discussed, as well as how they might integrate with other known pattern forming pathways known to modulate early developmental events (such as the Wnt/Ca2+pathway; [T.A. Westfall, B. Hjertos, D.C. Slusarski, Requirement for intracellular calcium modulation in zebrafish dorsal–ventral patterning, Dev. Biol. 259 (2003) 380–391]).
Keywords: Abbreviations; BP; Blastula Period; GP; Gastrula Period; MTs; microtubules; MFs; microfilamentsCa; 2+; Blastula Period; Gastrula Period; Zebrafish; Xenopus
Calcium signaling in plant cell organelles delimited by a double membrane by Tou-Cheu Xiong; Stéphane Bourque; David Lecourieux; Nicolas Amelot; Sabine Grat; Christian Brière; Christian Mazars; Alain Pugin; Raoul Ranjeva (pp. 1209-1215).
Increases in the concentration of free calcium in the cytosol are one of the general events that relay an external stimulus to the internal cellular machinery and allow eukaryotic organisms, including plants, to mount a specific biological response. Different lines of evidence have shown that other intracellular organelles contribute to the regulation of free calcium homeostasis in the cytosol. The vacuoles, the endoplasmic reticulum and the cell wall constitute storage compartments for mobilizable calcium. In contrast, the role of organelles surrounded by a double membrane (e.g. mitochondria, chloroplasts and nuclei) is more complex. Here, we review experimental data showing that these organelles harbor calcium-dependent biological processes. Mitochondria, chloroplasts as well as nuclei are equipped to generate calcium signal on their own. Changes in free calcium in a given organelle may also favor the relocalization of proteins and regulatory components and therefore have a profound influence on the integrated functioning of the cell. Studying, in time and space, the dynamics of different components of calcium signaling pathway will certainly give clues to understand the extraordinary flexibility of plants to respond to stimuli and mount adaptive responses. The availability of technical and biological resources should allow breaking new grounds by unveiling the contribution of signaling networks in integrative plant biology.
Keywords: Calcium; Plant cell signaling; Plant cell organization; Cell compartmentation; Dynamics of cytosolic and organelle calcium
New perspectives on the role of SERCA2's Ca2+ affinity in cardiac function by P. Vangheluwe; K.R. Sipido; L. Raeymaekers; F. Wuytack (pp. 1216-1228).
Cardiomyocyte relaxation and contraction are tightly controlled by the activity of the cardiac sarco(endo)plasmic reticulum (SR) Ca2+ transport ATPase (SERCA2a). The SR Ca2+-uptake activity not only determines the speed of Ca2+ removal during relaxation, but also the SR Ca2+ content and therefore the amount of Ca2+ released for cardiomyocyte contraction. The Ca2+ affinity is the major determinant of the pump's activity in the physiological Ca2+ concentration range. In the heart, the affinity of the pump for Ca2+ needs to be controlled between narrow borders, since an imbalanced affinity may evoke hypertrophic cardiomyopathy. Several small proteins (phospholamban, sarcolipin) adjust the Ca2+ affinity of the pump to the physiological needs of the cardiomyocyte. It is generally accepted that a chronically reduced Ca2+ affinity of the pump contributes to depressed SR Ca2+ handling in heart failure. Moreover, a persistently lower Ca2+ affinity is sufficient to impair cardiomyocyte SR Ca2+ handling and contractility inducing dilated cardiomyopathy in mice and humans. Conversely, the expression of SERCA2a, a pump with a lower Ca2+ affinity than the housekeeping isoform SERCA2b, is crucial to maintain normal cardiac function and growth. Novel findings demonstrated that a chronically increased Ca2+ affinity also may trigger cardiac hypertrophy in mice and humans. In addition, recent studies suggest that some models of heart failure are marked by a higher affinity of the pump for Ca2+, and hence by improved cardiomyocyte relaxation and contraction. Depressed cardiomyocyte SR Ca2+ uptake activity may therefore not be a universal hallmark of heart failure.
Keywords: SR Ca; 2+; ATPase; Phospholamban; Sarcolipin; Cardiac hypertrophy; Heart failure; SERCA2a
Neuronal Ca2+ signaling via caldendrin and calneurons by Marina Mikhaylova; Yogendra Sharma; Carsten Reissner; Falko Nagel; Penmatsa Aravind; Bheemreddy Rajini; Karl-Heinz Smalla; Eckart D. Gundelfinger; Michael R. Kreutz (pp. 1229-1237).
The calcium sensor protein caldendrin is abundantly expressed in neurons and is thought to play an important role in different aspects of synapto-dendritic Ca2+ signaling. Caldendrin is highly abundant in the postsynaptic density of a subset of excitatory synapses in brain and its distinct localization raises several decisive questions about its function. Previous work suggests that caldendrin is tightly associated with Ca2+- and Ca2+ release channels and might be involved in different aspects of the organization of the postsynaptic scaffold as well as with synapse-to-nucleus communication. In this report we introduce two new EF-hand calcium sensor proteins termed calneurons that apart from calmodulin represent the closest homologues of caldendrin in brain. Calneurons have a different EF-hand organization than other calcium sensor proteins, are prominently expressed in neurons and will presumably bind Ca2+ with higher affinity than caldendrin. Despite some significant structural differences it is conceivable that they are involved in similar Ca2+ regulated processes like caldendrin and neuronal calcium sensor proteins.
Keywords: Abbreviations; ANS; 8-Anilino-1-naphthalene sulfonic acid; Ca; 2+; Calcium; CaM; Calmodulin; ITC; Isothermal titration calorimetry; Mg; 2+; Magnesium; NCS; neuronal calcium sensorEF-hand; Ca; 2+; signaling; Caldendrin; Calmodulin; Calneurons; Ca; 2+; binding affinities; CaBP1; Structural modeling
Deciphering function and mechanism of calcium-binding proteins from their evolutionary imprints by Reginald O. Morgan; Silvia Martin-Almedina; Montserrat Garcia; Jorge Jhoncon-Kooyip; Maria-Pilar Fernandez (pp. 1238-1249).
Calcium-binding proteins regulate ion metabolism and vital signalling pathways in all living organisms. Our aim is to rationalize the molecular basis of their function by studying their evolution using computational biology techniques. Phylogenetic analysis is of primary importance for classifying cognate orthologs; profile hidden Markov models (HMM) of individual subfamilies discern functionally relevant sites by conservation probability analysis; and 3-dimensional structures display the integral protein in context. The major classifications of calcium-binding proteins, viz. EF-hand, C2 and ANX, exhibit structural diversity in their HMM fingerprints at the subfamily level, with functional consequences for protein conformation, exposure of receptor interaction sites and/or binding to membrane phospholipids. Calmodulin, S100 and annexin families were characterized in Petromyzon marinus (sea lamprey) to document genome duplication and gene creation events during the key evolutionary transition to primitive vertebrates. Novel annexins from diverse organisms revealed calcium-binding domains with accessory structural features that define their unique molecular fingerprints, protein interactivity and functional specificity. These include the first single-domain, bacterial annexin in Cytophaga hutchinsonii, the 21 tetrad annexins from the unicellular protist Giardia intestinalis, an ancestor to land plant annexins from the green alga Ostreococcus lucimarinus, invertebrate octad annexins and a critical polymorphism in human ANXA7. Receptor docking models supported the hypothesis of a potential interaction between annexin and C2 domains as a propitious mechanism for ensuring membrane translocation during signal transduction.
Keywords: Computational biology; Functional determinant; Hidden Markov model (HMM); Membrane-binding mechanism; Molecular evolution; Receptor docking
Use of a fluorescent polarization based high throughput assay to identify new Calmodulin ligands by Rania Dagher; Claire Pigault; Dominique Bonnet; Damien Boeglin; Christelle Pourbaix; Marie-Claude Kilhoffer; Pascal Villa; Camille G. Wermuth; Marcel Hibert; Jacques Haiech (pp. 1250-1255).
In order to develop a fluorescence polarization (FP) assay for calcium binding proteins, a fluorescent peptides based library of 1328 compounds has been synthesized. The use of this library has been validated by setting up a FP-high-throughput screening (FP-HTS) assay for calmodulin using the synthetic gene product (synCaM). With this assay, a set of 880 FDA approved compounds was screened. Besides the promazine class, we discovered two new classes of compounds that interact with calmodulin in a calcium dependent manner. One class has compounds with anti-histaminic/spasmolytic activities, and the other one are detergents with antibacterial activities.
Keywords: Fluorescence polarization/anisotropy; FP-High-throughput screening; synCaM; Fluorescent peptides based library
Characterization of the Munc13-calmodulin interaction by photoaffinity labeling by Kalina Dimova; Hiroshi Kawabe; Andrea Betz; Nils Brose; Olaf Jahn (pp. 1256-1265).
Sensing of and response to transient increases in the residual presynaptic Ca2+ levels are important adaptive mechanisms that define the short-term plasticity characteristics of neurons. Due to their essential function in synaptic vesicle priming and in the modulation of synaptic strength, Munc13 proteins have emerged as key regulators of these adaptive mechanisms. Indeed, Munc13-1 and ubMunc13-2 contain a conserved calmodulin (CaM) binding site and the Ca2+-dependent interaction of these Munc13 isoforms with CaM constitutes a molecular mechanism that transduces residual Ca2+ signaling to the synaptic exocytotic machinery. Here, we used Munc13-derived model peptides in photoaffinity labeling (PAL) experiments to demonstrate the stoichiometric and Ca2+-dependent CaM binding of the other members of the Munc13 family, bMunc13-2 and Munc13-3, via structurally distinct non-conserved binding sites. A PAL-based Ca2+ titration assay revealed that all Munc13 isoforms can form a complex with CaM already at low Ca2+ concentrations just above resting levels, underscoring the Ca2+ sensor/effector function of this interaction in short-term synaptic plasticity phenomena.
Keywords: Vesicle priming; Munc13; Calmodulin; Short-term plasticity; Ca; 2+; sensitivity; Photoaffinity labeling
Development of a novel bioavailable inhibitor of the calmodulin-regulated protein kinase MLCK: A lead compound that attenuates vascular leak by Heather A. Behanna; D. Martin Watterson; Hantamalala Ralay Ranaivo (pp. 1266-1274).
Tissue barriers involving epithelial and endothelial cell layers are critical to homeostasis, regulating passage of water, macromolecules, cells and certain classes of small molecules via two distinct cellular mechanisms, transcellular or paracellular. Endothelial or epithelial barrier dysfunction is a key component of pathophysiology in diverse diseases and injuries that have a broad impact on survival and quality of life. However, effective and safe small molecule therapeutics for these disorders are lacking. Success in development would therefore fill a major unmet medical need across multiple disease areas. Myosin light chain kinase (MLCK), a highly specialized calcium/calmodulin (CaM) regulated protein kinase, modulates barrier function through its regulation of intracellular contractile processes. MLCK levels and activity are increased in various animal models of disease and in human clinical disease samples. Our prior work with a genetic knockout (KO) mouse strain for the long form of MLCK, MLCK210, has identified MLCK as a drug discovery target for endothelial and epithelial barrier dysfunction. We describe here the development of a selective, bioavailable, stable inhibitor of MLCK that attenuates barrier dysfunction in mice comparable to that seen with the MLCK KO mice. The inhibitor compound 6 is stable in human microsomal metabolic stability assays and can be synthesized in a high-yielding and facile synthetic process. These results provide a foundation for and demonstrate the feasibility of future medicinal chemistry refinement studies directed toward the development of novel therapies for disorders involving barrier dysfunction.
Keywords: Myosin light chain kinase; Barrier dysfunction; Genetic knockout; Inflammatory disorder; Drug discovery; Structure–activity relationship
Nuclear Ca2+/calmodulin-dependent protein kinase II in the murine heart by Bailing Li; John R. Dedman; Marcia A. Kaetzel (pp. 1275-1281).
Ca2+ signaling through CaMKII is critical in regulating myocyte function with regard to excitation–contraction–relaxation cycles and excitation–transcription coupling. To investigate the role of nuclear CaMKII in cardiac function, transgenic mice were designed and generated to target the expression of a CaMKII inhibitory peptide, AIP (KKALRRQEAVDAL), to the nucleus. The transgenic construct consists of the murine α-myosin heavy chain promoter followed by the expression unit containing nucleotides encoding a four repeat concatemer of AIP (AIP4) and a nuclear localization signal (NLS). Western blot and immunohistochemical analyses demonstrate that AIP4 is expressed only in the nucleus of cardiac myocytes of the transgenic mice (NLS-AIP4). The function of cytoplasmic CaMKII is not affected by the expression of AIP4 in the nucleus. Inhibition of nuclear CaMKII activity resulted in reduced translocation of HDAC5 from nucleus to cytoplasm in NLS-AIP4 mouse hearts. Loss of nuclear CaMKII activity causes NLS-AIP4 mice to have smaller hearts than their nontransgenic littermates. Transcription factors including CREB and NFκB are not regulated by cardiac nuclear CaMKII. With physiological stresses such as pregnancy or aging (8 months), NLS-AIP4 mice develop hypertrophy symptoms including enlarged atria, systemic edema, sedentariness, and morbidity. RT-PCR analyses revealed that the hypertrophic marker genes, such as ANF and β-myosin heavy chain, were upregulated in pregnancy stressed mice. Our results suggest that absence of adequate Ca2+ signaling through nuclear CaMKII regulated pathways leads to development of cardiac disease.
Keywords: Nuclear CaMKII; Ca; 2+; Hypertrophy; Inhibitor peptide; Heart; Murine
An update of the S100 nomenclature by Ingo Marenholz; Ruth C. Lovering; Claus W. Heizmann (pp. 1282-1283).
The plethora of names given to S100 proteins resulted in considerable confusion. Here we present the official and updated nomenclature of this protein family, approved by the HGNC (HUGO gene nomenclature committee) and ECS (European Calcium Society).
Keywords: S100 protein; Nomenclature; Calcium; Chromosomal localization
Recognition of the tumor suppressor protein p53 and other protein targets by the calcium-binding protein S100B by Paul T. Wilder; Jing Lin; Catherine L. Bair; Thomas H. Charpentier; Dong Yang; Melissa Liriano; Kristen M. Varney; Andrew Lee; Amos B. Oppenheim; Sankar Adhya; France Carrier; David J. Weber (pp. 1284-1297).
S100B is an EF-hand containing calcium-binding protein of the S100 protein family that exerts its biological effect by binding and affecting various target proteins. A consensus sequence for S100B target proteins was published as (K/R)(L/I)xWxxIL and matches a region in the actin capping protein CapZ (V.V. Ivanenkov, G.A. Jamieson, Jr., E. Gruenstein, R.V. Dimlich, Characterization of S-100b binding epitopes. Identification of a novel target, the actin capping protein, CapZ, J. Biol. Chem. 270 (1995) 14651–14658). Several additional S100B targets are known including p53, a nuclear Dbf2 related (NDR) kinase, the RAGE receptor, neuromodulin, protein kinase C, and others. Examining the binding sites of such targets and new protein sequence searches provided additional potential target proteins for S100B including Hdm2 and Hdm4, which were both found to bind S100B in a calcium-dependent manner. The interaction between S100B and the Hdm2 and/or the Hdm4 proteins may be important physiologically in light of evidence that like Hdm2, S100B also contributes to lowering protein levels of the tumor suppressor protein, p53. For the S100B–p53 interaction, it was found that phosphorylation of specific serine and/or threonine residues reduces the affinity of the S100B–p53 interaction by as much as an order of magnitude, and is important for protecting p53 from S100B-dependent down-regulation, a scenario that is similar to what is found for the Hdm2–p53 complex.
Keywords: S100B; Calcium-binding protein; S100 protein; p53; Hdm2; Mdm2
Biophysical characterization of S100A8 and S100A9 in the absence and presence of bivalent cations by Thomas Vogl; Nadja Leukert; Katarzyna Barczyk; Kerstin Strupat; Johannes Roth (pp. 1298-1306).
S100A8 and S100A9 are two proinflammatory molecules belonging to the S100 family of calcium-binding proteins. Common to all S100 proteins S100A8 and S100A9 form non-covalently associated complexes which have been shown to exhibit different functional properties. Besides dimerization, recent research is focused on the importance of higher oligomeric structures of S100 proteins induced by bivalent cations. While S100A8/S100A9-heterodimers are formed in the absence of calcium, tetramerization is strictly calcium-dependent. Heterodimer formation is not a simple process and our biophysical analyses (FRET, ESI-MS) demonstrate that simply mixing both subunits is not sufficient to induce complex formation. Steps of denaturation/renaturation are necessary for the recombinant complex to show identical biophysical properties as S100A8/S100A9 obtained from granulocytes. In addition to calcium both proteins are able to bind zinc with high affinity. Here we demonstrate for the first time by different biophysical methods (MALDI-MS, ESI-MS, fluorescence spectroscopy) that zinc-binding, like calcium, induces (S100A8/S100A9)2-tetramers. Using mass spectrometric investigations we demonstrate that zinc triggers the formation of (S100A8/S100A9)2-tetramers by zinc-specific binding sites rather than by interactions with calcium-specific EF-hands. The zinc-induced tetramer is structurally very similar to the calcium-induced tetramer. Thus, like calcium, zinc acts as a regulatory factor in S100A8/S100A9-dependent signaling pathways.
Keywords: Calcium-binding proteins; MRP8; MRP14; Calgranulin; Calprotectin; Complex formation; Calcium; Zinc
S100A1-deficient male mice exhibit increased exploratory activity and reduced anxiety-related responses by Gabriele E. Ackermann; Ingo Marenholz; David P. Wolfer; Wood Yee Chan; Beat Schäfer; Paul Erne; Claus W. Heizmann (pp. 1307-1319).
S100 proteins comprise a family of Ca2+ binding proteins of at least 21 members. They are distinctly expressed in a variety of cell types and tissues and are thought to play unique roles, although they share a high degree of sequence homology and expression overlap. S100A1 is prominently expressed in the heart, where it takes part in Ca2+-cycling. Its role in the central nervous system (CNS) is largely unknown. We have generated S100A1-deficient mice by gene trap mutagenesis to study the involvement of S100A1 in the cytoarchitecture of the brain, in learning and memory, and in avoidance-approach behavior. S100A1 knock out (KO) mice develop well and their brains present with normal morphology. In wild type (Wt) mice, S100A1 protein was found in the hippocampus, cerebral cortex and amygdala, and partially co-localized with the astrocyte marker glial fibrillary acidic protein (GFAP) in the stratum radiatum of the hippocampus. Astrocytes and neurons of S100A1KO mice did not differ from those of Wt mice regarding shape, distribution and density. In the water maze, S100A1KO mice performed equally well as Wt, implying that S100A1 is not involved in spatial learning and memory. In avoidance-approach tests, predominantly male S100A1KO mice showed reduced anxiety-like responses and enhanced explorative activities. We conclude that S100A1 plays a role in modulating innate fear and exploration of novel stimuli.
Keywords: Abbreviations; CNS; central nervous system; HPLC; high performance liquid chromatography; KO; knock out; PGK; phosphoglycerate kinase-1; PCR; polymerase chain reaction; RT; reverse transcription; Wt; wild typeS100; Mouse; CNS; Anxiety; Exploration; Immunohistochemistry
Proteolytic cleavage of annexin 1 by human leukocyte elastase by Ursula Rescher; Verena Goebeler; Andreas Wilbers; Volker Gerke (pp. 1320-1324).
Annexin 1 has been shown to participate through its unique N-terminal domain in the recruitment and activation of leukocytes at sites of inflammation. Peptides derived from this domain are true mimetics of the annexin 1 action in all inflammation models tested and most likely serve as the active entities generated at sites of inflammation. To elucidate mechanisms underlying peptide generation we used isolated blood leukocytes and endothelial cell monolayers. We show that following endothelial adhesion, annexin 1 was externalized from leukocytes and rapidly cleaved. Addition of purified annexin 1 to degranulating leukocytes resulted in the truncation of annexin 1, which seemed to depend on the proteolytic activity of human leukocyte elastase (HLE). The capacity of elastase to proteolytically cleave annexin 1 was confirmed using both purified annexin 1 and HLE. The identification of annexin 1 as a substrate for HLE supports the model in which annexin 1 participates in regulating leukocyte emigration into inflamed tissue through N-terminal peptides generated at inflammatory sites.
Keywords: Calcium; Endothelial cell; Inflammation; Elastase
Annexin A2 recognises a specific region in the 3′-UTR of its cognate messenger RNA by Hanne Hollås; Ingvild Aukrust; Stine Grimmer; Elin Strand; Torgeir Flatmark; Anni Vedeler (pp. 1325-1334).
Annexin A2 is a multifunctional Ca2+- and lipid-binding protein. We previously showed that a distinct pool of cellular Annexin A2 associates with mRNP complexes or polysomes associated with the cytoskeleton. Here we report in vitro and in vivo experiments showing that Annexin A2 present in this subset of mRNP complexes interacts with its cognate mRNA and c- myc mRNA, but not with β2 -microglobulin mRNA translated on membrane-bound polysomes. The protein recognises sequence elements within the untranslated regions, but not within the coding region, of its cognate mRNA. Alignment of the Annexin A2-binding 3′-untranslated regions of annexin A2 mRNA from several species reveals a five nucleotide consensus sequence 5′-AA(C/G)(A/U)G. The Annexin A2-interacting region of the 3′-untranslated region can be mapped to a sequence of about 100 nucleotides containing two repeats of the consensus sequence. The binding elements appear to involve both single and double stranded regions, indicating that a specific higher order mRNA structure is required for binding to Annexin A2. We suggest that this type of interaction is representative for a group of mRNAs translated on cytoskeleton-bound polysomes.
Keywords: Abbreviations; AnxA2; Annexin A2 protein; anx; A2; annexin A2 mRNA or cDNA; AnxA2t; Annexin A2 heterotetramer (AnxA2; 2; p11; 2; ); BSA; bovine serum albumin; CBP; cytoskeleton-bound polysomes; CDR; coding region; ds; double stranded; FL; full-length; FP; free polysomes; HC; heavy chain; MBP; membrane-bound polysomes; mRNP; messenger ribonucleotide particle; nt; nucleotides; PCR; polymerase chain reaction; RAM; rabbit anti-mouse IgG; RT-PCR; reverse trancriptase PCR; ss; single stranded; TEV; Tobacco Etch Virus; UTR; untranslated regionAnnexin A2; mRNA; 3′-UTR; mRNA-binding; consensus sequence; c-; myc
Nuclear translocation of the calcium-binding protein ALG-2 induced by the RNA-binding protein RBM22 by P. Montaville; Y. Dai; C.Y. Cheung; K. Giller; S. Becker; M. Michalak; S.E. Webb; A.L. Miller; J. Krebs (pp. 1335-1343).
By yeast two-hybrid screening using the calcium-binding protein ALG-2 as bait a new target of ALG-2 was identified, the RNA-binding protein RBM22. In order to confirm these interactions in vivo we prepared fluorescent constructs by using the monomeric red fluorescent protein to label ALG-2 and the enhanced green fluorescent protein to label RBM22. Confocal microscopy of NIH 3T3 cells transfected with either ALG-2 or RBM22 expression constructs encoding fluorescent fusion proteins alone revealed that the majority of ALG-2 was localized in the cytoplasm whereas RBM22 was located in the nucleus. When cells were co-transfected with expression vectors encoding both fusion proteins ALG-2 was found in the nucleus indicating that RBM22 which can shuttle between the cytoplasm and the nucleus may play a role in nuclear translocation of ALG-2. Using zebrafish as a model mRNA homologues of ALG-2 and RBM22 were microinjected into the blastodisc-yolk margin of zebrafish embryos at the 1-cell stage followed by monitoring the fusion proteins during development of the zebrafish. Hereby, we observed that ALG-2 alone evenly distributed within the cell, whereas in the presence of RBM22 the two proteins co-localized within the nucleus. More than 95% of the two proteins co-localized within the same area in the nucleus suggesting a functional interaction between the Ca2+-signaling protein ALG-2 and the RNA-binding protein RBM22.
Keywords: ALG-2, RBM22; Ca; 2+; -binding protein; RNA-binding protein; Zebrafish development; Confocal microscopy
Cytobiological consequences of calcium-signaling alterations induced by human viral proteins by Mounia Chami; Bénédicte Oulès; Patrizia Paterlini-Bréchot (pp. 1344-1362).
Since calcium-signaling regulates specific and fundamental cellular processes, it represents the ideal target of viral proteins, in order for the virus to control cellular functions and favour its persistence, multiplication and spread. A detailed analysis of reports focused on the impact of viral proteins on calcium-signaling has shown that virus-related elevations of cytosolic calcium levels allow increased viral protein expression (HIV-1, HSV-1/2), viral replication (HBx, enterovirus 2B, HTLV-1 p12I, HHV-8, EBV), viral maturation (rotavirus), viral release (enterovirus 2B) and cell immortalization (EBV). Interestingly, virus-induced decreased cytosolic calcium levels have been found to be associated with inhibition of immune cells functions (HIV-1 Tat, HHV-8 K15, EBV LMP2A). Finally, several viral proteins are able to modulate intracellular calcium-signaling to control cell viability (HIV-1 Tat, HTLV-1 p13II, HCV core, HBx, enterovirus 2B, HHV-8 K7). These data point out calcium-signaling as a key cellular target for viral infection and should stimulate further studies exploring new calcium-related therapeutic strategies.
Keywords: Abbreviations; aa; amino acids; [Ca; 2+; ]; cyt; cytosolic calcium-concentration; [Ca; 2+; ]; er; endoplasmic reticulum calcium-concentration; [Ca; 2+; ]; Golgi; Golgi apparatus calcium-concentration; [Ca; 2+; ]; mt; mitochondrial calcium-concentration; Ca; 2+; calcium; EBV; Epstein–Barr virus; ER; endoplasmic reticulum; HBV; hepatitis B virus; HCV; hepatitis C virus; HHV-8; human herpesvirus type-8; HIV-1; human immunodeficiency virus type-1; HPV; human papillomavirus; HSV-1/2; herpes simplex virus type-1 and type-2; HTLV-1; human T lymphotropic virus type-1; IP; 3; inositol 1,4,5-triphosphate; IP; 3; R; inositol triphosphate receptor; kb; kilobase; kDa; kilodalton; NCX/HCX; Na; +; /Ca; 2+; exchanger and H; +; /Ca; 2+; exchanger; NFAT; nuclear factor of activated T cell; PMCA; plasma membrane Ca; 2+; -ATPase; SERCA; sarco-Endoplasmic reticulum Ca; 2+; -ATPase; ROC; receptor operated channel; RYR; ryanodine receptor; SERCA; sarco-Endoplasmic reticulum Ca; 2+; -ATPase; SOC; store operated channel; SPCA; secretory pathway Ca; 2+; -ATPase; TRPC; members of the transient receptor potential ion-channel familyCalcium-signaling; Human virus; Viral protein; Signal transduction; Viral infection; Calcium measurement