Thermal stability of nacre proteins of the polynesian pearl oyster: a proteomic study.

Mollusc shells are organic-inorganic composites that are often preserved in the fossil record. However, the way the organic fraction, also called shell matrix, gets fossilized remains an unsolved question, in spite of several old and more recent studies. In the present paper, we have tried to mimic a diagenetic process by constantly heating for ten days at 100°C fresh nacre powder samples of the Polynesian pearl oyster Pinctadamargaritifera. Each day, aliquots of nacre powder were sampled and the matrix was subsequently extracted. It was further analysed by direct weigh quantification, by immunological techniques and by proteomics. Our preliminary data suggest that nacre proteins, when heat…

The skeletal proteome of the coral Acropora millepora: the evolution of calcification by co-option and domain shuffling.

14 pages; International audience; In corals, biocalcification is a major function that may be drastically affected by ocean acidification (OA). Scleractinian corals grow by building up aragonitic exoskeletons that provide support and protection for soft tissues. Although this process has been extensively studied, the molecular basis of biocalcification is poorly understood. Notably lacking is a comprehensive catalog of the skeleton-occluded proteins-the skeletal organic matrix proteins (SOMPs) that are thought to regulate the mineral deposition. Using a combination of proteomics and transcriptomics, we report the first survey of such proteins in the staghorn coral Acropora millepora. The or…

The Representation of Shopping in Children’s Books

Purpose The purpose of this paper is to identify the representations, figures and processes of shopping/commerce in books published in France that are aimed at three to seven-year-olds. Design/methodology/approach A semiotic analysis of nearly 50 books published over the past 60 years. Findings These books reveal a broad diversity in the images of shops given to children (ranging from the traditional shop, a source of pleasure and creator of social ties, to the hypermarket/megastore, a symbol of stress and overconsumption) and the wealth of information that is given to children to help them assimilate the process of a shopping transaction. Originality/value The originality and richness of …

The skeleton of the staghorn coral Acropora millepora: molecular and structural characterization.

15 pages; International audience; The scleractinian coral Acropora millepora is one of the most studied species from the Great Barrier Reef. This species has been used to understand evolutionary, immune and developmental processes in cnidarians. It has also been subject of several ecological studies in order to elucidate reef responses to environmental changes such as temperature rise and ocean acidification (OA). In these contexts, several nucleic acid resources were made available. When combined to a recent proteomic analysis of the coral skeletal organic matrix (SOM), they enabled the identification of several skeletal matrix proteins, making A. millepora into an emerging model for biomi…

Shell repair in the clam Ruditapes philippinarum, affected by the Brown Ring Disease (BRD): a biochemical and serological study.

AbstractFor more than two decades, the edible clam Ruditapes philippinarum has been affected by the Brown Ring Disease (BRD), a bacterial infection characterized by the formation of a brown organic deposit in the internal side of the valves. Although this infection is often lethal, in some cases specimens can overcome it by remineralizing over the organic deposit. The goal of the present study is to compare biochemically and immunologically the shell matrices of repaired and healthy zones. Our data suggest that the repair zones exhibit certain variability, which would be the direct consequence of a modification of the secretory regime of calcifying tissues responsible of the repair process.

The shell organic matrix of the crossed lamellar queen conch shell (Strombus gigas)

10 pages; International audience; In molluscs, the shell organic matrix comprises a large set of biomineral-occluded proteins, glycoproteins and polysaccharides that are secreted by the calcifying mantle epithelium, and are supposed to display several functions related to the synthesis of the shell. In the present paper, we have characterized biochemically the shell matrix associated to the crossed-lamellar structure of the giant queen conch Strombus gigas. The acid-soluble (ASM) and acid-insoluble (AIM) matrices represent an extremely minor fraction of the shell. Both are constituted of polydisperse and of few discrete proteins among which three fractions, obtained by preparative SDS-PAGE …

The shell-forming proteome of Lottia gigantea reveals both deep conservations and lineage-specific novelties

19 pages; International audience; Proteins that are occluded within the molluscan shell, the so-called shell matrix proteins (SMPs), are an assemblage of biomolecules attractive to study for several reasons. They increase the fracture resistance of the shell by several orders of magnitude, determine the polymorph of CaCO(3) deposited, and regulate crystal nucleation, growth initiation and termination. In addition, they are thought to control the shell microstructures. Understanding how these proteins have evolved is also likely to provide deep insight into events that supported the diversification and expansion of metazoan life during the Cambrian radiation 543 million years ago. Here, we p…

Variability of shell repair in the Manila clam Ruditapes philippinarum affected by the Brown Ring Disease: A microstructural and biochemical study

11 pages; International audience; For more than two decades, the Manila clam Ruditapes philippinarum has been regularly affected by Brown Ring Disease (BRD), an epizootic event caused by the bacterium Vibrio tapetis and characterized by the development of a brown deposit on the inner face of valves. Although BRD infection is often lethal, some clams recover by mineralizing a new repair shell layer, which covers the brown deposit and fully isolates it from living tissues. In order to understand this specific shell repair process, the microstructures of repaired zones were compared to those of shells unaffected by BRD. In addition, the organic matrix associated with unaffected shells and to r…

Spine and test skeletal matrices of the Mediterranean sea urchinArbacia lixula- a comparative characterization of their sugar signature

15 pages; International audience; Calcified structures of sea urchins are biocomposite materials that comprise a minor fraction of organic macromolecules, such as proteins, glycoproteins and polysaccharides. These macromolecules are thought to collectively regulate mineral deposition during the process of calcification. When occluded, they modify the properties of the mineral. In the present study, the organic matrices (both soluble and insoluble in acetic acid) of spines and tests from the Mediterranean black sea urchin Arbacia lixula were extracted and characterized, in order to determine whether they exhibit similar biochemical signatures. Bulk characterizations were performed by mono-di…

Biochemical Characterization of the Soluble Organic Matrix of Gastroliths from Decapods.

10 pages; International audience; Gastroliths are disc-shaped calcium storage structures cyclically elaborated by some decapods in their stomach wall. In the present study, we extracted the gastrolith organic matrix from three different decapods, two crayfishes and one lobster, and began to characterize the soluble matrix obtained after decalcification by acetic acid. By using different staining (silver nitrate, Coomassie Blue, Stains-all) after SDS-PAGE, we evidenced that these soluble fractions are rich in polypeptides, some seems common, others are completely different. Among them, putative calcium binding proteins might be present and have to be further characterized. This soluble organ…

The impact of self-esteem on the relationship between the endorser and the consumers’s behavioural intentions

Titre du congrès : «Influencing consumers' behaviors - 'Art nouveau' in consumer research»; International audience; Using endorsers to promote products is a popular advertising strategy. Five major types of endorsers used in advertising are: celebrities, typical consumers, professional experts, company presidents and models. To date, most academic research on the endorser’s effectiveness has focused on celebrity endorsement. The objective of this study is to expand on the other endorsement types literature by studying the effectiveness of advertising using five types of endorsers with incorporating consumers’ self‐esteem. Results suggest that this variable is a moderator of the direct effec…

Caspartin: thermal stability and occurrence in mollusk calcified tissues.

8 pages; International audience; The fan mussel Pinna nobilis secretes a two-layered shell, the external layer of which is composed of long calcitic prisms. The dissolution of these prisms releases an assemblage of acidic intracrystalline proteins, among which caspartin, an Asp-rich protein (Marin et al., 2005). A polyclonal antibody raised against purified caspartin was used for estimating its thermal stability, and for investigating the presence of cross-reacting proteins in the shell matrices of several mollusks. In the first case, prisms of Pinna nobilis were heated at 100°C, for one to eleven days. The degradation of caspartin was followed on SDS-PAGE and on Western-blot. The experimen…

The test skeletal matrix of the black sea urchin Arbacia lixula

11 pages; International audience; In the field of biomineralization, the past decade has been marked by the increasing use of high throughput techniques, i.e. proteomics, for identifying in one shot the protein content of complex macromolecular mixtures extracted from mineralized tissues. Although crowned with success, this approach has been restricted so far to a limited set of key-organisms, such as the purple sea urchin Strongylocentrotus purpuratus, the pearl oyster or the abalone, leaving in the shadow non-model organisms. As a consequence, it is still unknown to what extent the calcifying repertoire varies, from group to group, at high (phylum, class), median (order, family) or low (g…

A minimal molecular toolkit for mineral deposition? Biochemistry and proteomics of the test matrix of adult specimens of the sea urchin Paracentrotus lividus.

12 pages; International audience; The sea urchin endoskeleton consists of a magnesium-rich biocalcite comprising a small amount of occluded organic macromolecules. This structure constitutes a key-model for understanding the mineral - organics interplay, and for conceiving in vitro bio-inspired materials with tailored properties. Here we employed a deep-clean technique to purify the occluded proteins from adult Paracentrotus lividus tests. We characterized them by 1- and 2D-electrophoreses, ELISA and immunoblotting, and using liquid chromatography coupled with Mass Spectrometry (nanoLC-MS/MS), we identified two metalloenzymes (carbonic anhydrase and MMP), a set of MSP130 family members, sev…

Nacre calcification in the freshwater mussel Unio pictorum: carbonic anhydrase activity and purification of a 95 kDa calcium-binding glycoprotein.

9 pages; International audience; The formation of the molluscan shell is finely tuned by macromolecules of the shell organic matrix. Previous results have shown that the acid-soluble fraction of the nacre matrix of the freshwater paleoheterodont bivalve Unio pictorum shell displays a number of remarkable properties, such as calcium-binding activity, the presence of extensive glycosylations and the capacity to interfere at low concentration with in vitro calcium carbonate precipitation. Here we have found that the nacre-soluble matrix exhibits a carbonic anhydrase activity, an important function in calcification processes. This matrix is composed of three main proteinaceous discrete fraction…

Shell matrices of recent rhynchonelliform brachiopods: microstructures and glycosylation studies.

ABSTRACTLike most metazoan biomineralisations, the brachiopod shell is the end product of a biologically controlled calcification process. The main agent of the control is the extracellular matrix, which is secreted by the outer mantle epithelium. This matrix mediates the calcification process by allowing crystal nucleation and elongation in specific orientations and finally, by stopping crystal growth. The proteinaceous moiety of brachiopod shell matrices has been extensively studied. Less known are the post-translational modifications that occur in these matrices, in particular glycosylations. In this comparison of five species of Recent articulated brachiopods, the ratio of soluble to in…

Facilitated transport of diuron and glyphosate in high copper vineyard soils.

6 pages; International audience; The fate of organic herbicides applied to agricultural fields may be affected by other soil amendments, such as copper applied as a fungicide. The effect of copper on the leaching of diuron and glyphosate through a granitic and a calcareous soil was studied in the laboratory using sieved-soil columns. Each soil was enriched with copper sulfate to obtain soil copper concentrations of 125, 250, 500, and 1000 mg kg(-1). Glyphosate leaching was influenced by soil pH and copper concentration, whereas diuron leaching was not. In the calcareous soil, glyphosate leaching decreased as copper levels increased from 17 mg kg(-1) (background) to 500 mg kg(-1). In the gra…

Identification of Two Carbonic Anhydrases in the Mantle of the European Abalone Haliotis tuberculata (Gastropoda, Haliotidae): Phylogenetic Implications

Carbonic anhydrases (CAs) represent a diversified family of metalloenzymes that reversibly catalyze the hydration of carbon dioxide. They are involved in a wide range of functions, among which is the formation of CaCO(3) skeletons in metazoans. In the shell-forming mantle tissues of mollusks, the location of the CA catalytic activity is elusive and gives birth to contradicting views. In the present paper, using the European abalone Haliotis tuberculata, a key model gastropod in biomineralization studies, we identified and characterized two CAs (htCA1 and htCA2) that are specific of the shell-forming mantle tissue. We analyzed them in a phylogenetic context. Combining various approaches, inc…

Biochemical characteristics of the shell soluble organic matrix of some recent Rhynchonelliformea (Brachiopoda).

11 pages; International audience; To build up their shell, brachiopods secrete a mixture of proteins and polysaccharides, collectively called the shell organic matrix. This matrix is supposed to mediate the calcification process by allowing the crystal nucleation and elongation and finally, by stopping the crystal growth. The aim of this work is, firstly, to observe at nanoscale the close relationships between the mineral and the organic phases, and secondly, to characterize the acetic acid-soluble matrices extracted from different articulate brachiopods. Recent articulate brachiopod shells were observed according to a standard procedure, which emphasizes the microstructural details. For th…

Metazoan calcium carbonate biomineralizations: macroevolutionary trends – challenges for the coming decade

AbstractCalcium carbonate-based biominerals, also referred as biocalcifications, are the most abundant biogenic mineralized products at the surface of the Earth. In this paper, we summarize general concepts on biocalcifications and we sketch macro-evolutionary trends throughout the history of the Earth, from Archean to Phanerozoic times. Then, we expose five fundamental issues that represent key-challenges in biocalcification researches for the coming decade: the first one concerns the comprehension of the micro- and nano-structure of calcium carbonate biominerals from a mineral viewpoint, while the second one deals with the understanding of the dynamic process of their fabrication. The thi…

La relation à la marque du Z consommateur : une étude exploratoire des causes de détachement

Calcification in the Shell of the Freshwater Bivalve Unio pictorum.

8 pages; International audience; The scope of the present study is to elucidate the primary structure of the shell proteins of the mollusk Unio pictorum. This freshwater bivalve exhibits an aragonitic shell with nacro-prismatic microstructures. The acetic acid-soluble matrix of the nacreous layer has been extracted, and analyzed on standard polyacrylamide gel under denaturing conditions. Five main proteins can be observed. Their respective apparent molecular weight is 95, 50, 29, 14 and 10 kDa. The three high molecular weight bands are acidic, glycosylated and bind calcium in vitro. These proteins are purified by preparative SDSPAGE. In addition, these fractions are tested to check their ab…

The shell matrix of the pulmonate land snail Helix aspersa maxima.

12 pages; International audience; In mollusks, the shell mineralization process is controlled by an array of proteins, glycoproteins and polysaccharides that collectively constitute the shell matrix. In spite of numerous researches, the shell protein content of a limited number of model species has been investigated. This paper presents biochemical data on the common edible land snail Helix aspersa maxima, a model organism for ecotoxicological purposes, which has however been poorly investigated from a biomineralization viewpoint. The shell matrix of this species was extracted and analyzed biochemically for functional in vitro inhibition assay, for amino acid and monosaccharides composition…

'Shellome': Proteins involved in mollusk shell biomineralization - diversity, functions.

18 pages; International audience

Caspartin and calprismin, two proteins of the shell calcitic prisms of the Mediterranean fan mussel Pinna nobilis.

We used the combination of preparative electrophoresis and immunological detection to isolate two new proteins from the shell calcitic prisms of Pinna nobilis, the Mediterranean fan mussel. The amino acid composition of these proteins was determined. Both proteins are soluble, intracrystalline, and acidic. The 38-kDa protein is glycosylated; the 17-kDa one is not. Ala, Asx, Thr, and Pro represent the dominant residues of the 38-kDa protein, named calprismin. An N-terminal sequence was obtained from calprismin. This sequence, which comprises a pattern of 4 cysteine residues, is not related to any known protein. The second protein, named caspartin, exhibits an unusual amino acid composition, …

The shell matrix of the freshwater mussel Unio pictorum (Paleoheterodonta, Unionoida). Involvement of acidic polysaccharides from glycoproteins in nacre mineralization.

13 pages; International audience; Among molluscs, the shell biomineralization process is controlled by a set of extracellular macromolecular components secreted by the calcifying mantle. In spite of several studies, these components are mainly known in bivalves from only few members of pteriomorph groups. In the present case, we investigated the biochemical properties of the aragonitic shell of the freshwater bivalve Unio pictorum (Paleoheterodonta, Unionoida). Analysis of the amino acid composition reveals a high amount of glycine, aspartate and alanine in the acid-soluble extract, whereas the acid-insoluble one is rich in alanine and glycine. Monosaccharidic analysis indicates that the in…

Comparative ultrastructure and carbohydrate composition of gastroliths from Astacidae, Cambaridae and Parastacidae freshwater crayfish (Crustacea, Decapoda)

21 pages; International audience; Crustaceans have to cyclically replace their rigid exoskeleton in order to grow. Most of them harden this skeleton by a calcification process. Some decapods (land crabs, lobsters and crayfish) elaborate calcium storage structures as a reservoir of calcium ions in their stomach wall, as so-called gastroliths. For a better understanding of the cyclic elaboration of these calcium deposits, we studied the ultrastructure of gastroliths from freshwater crayfish by using a combination of microscopic and physical techniques. Because sugars are also molecules putatively involved in the elaboration process of these biomineralizations, we also determined their carbohy…

Synthesis of calcium carbonate biological materials: how many proteins are needed?

In Nature, calcium carbonate biomineralizations are the most abundant mineralized structures of biological origin. Because many exhibit remarkable characteristics, several attempts have been made to use them as substitution materials for bone reconstruction or as models for generating biomimetic composites that exhibit tailored properties. CaCO3biomineralizations contain small amounts of amalgamate of proteins and polysaccharides that are secreted during the calcification process. They contribute to control the morphology of the crystallites and to spatially organize them in well-defined microstructures. These macromolecules, collectively defined as the skeletal matrix, have been the focus …

Diuron mobility through vineyard soils contaminated with copper

International audience; The herbicide diuron is frequently applied to vineyard soils in Burgundy, along with repeated treatments with Bordeaux mixture (a blend of copper sulfate and calcium hydroxide) that result in elevated copper concentrations. Cu could in principle affect the fate and transport of diuron or its metabolites in the soil either directly by complexation or indirectly by altering the populations or activity of microbes involved in their degradation. To assess the effect of high Cu concentrations on diuron transport, an experiment was designed with ten undisturbed columns of calcareous and acidic soils contaminated with 17–509 mg kg−1 total Cu (field-applied). Grass was plant…

Novel molluskan biomineralization proteins retrieved from proteomics: a case study with upsalin.

12 pages; International audience; The formation of the molluskan shell is regulated by an array of extracellular proteins secreted by the calcifying epithelial cells of the mantle. These proteins remain occluded within the recently formed biominerals. To date, many shell proteins have been retrieved, but only a few of them, such as nacreins, have clearly identified functions. In this particular case, by combining molecular biology and biochemical approaches, we performed the molecular characterization of a novel protein that we named Upsalin, associated with the nacreous shell of the freshwater mussel Unio pictorum. The full sequence of the upsalin transcript was obtained by RT-PCR and 5'/3…

Characterization of crustacyanin-A2 subunit as a component of the organic matrix of gastroliths from the crayfish Cherax quadricarinatus.

AbstractLike the lobsters, some terrestrial crabs and other crayfishes, the Australian red claw crayfish, Cherax quadricarinatus, elaborates in its stomach wall calcium storage structures called gastroliths. For understanding the cyclic elaboration and stabilization of these amorphous calcified structures, we studied the organic matrix (OM) of these paired biomineralizations. After decalcification with acetic acid, we analysed the proteinaceous components of an acetic acid-insoluble fraction by two-dimensional electrophoresis. Nine spots were digested by trpsin and the tryptic peptides were sequenced by nanoLC-nanoESI-MS/MS mass spectrometry. About 100 peptidic sequences were compared to se…

Shell proteome of rhynchonelliform brachiopods.

7 pages; International audience; Brachiopods are a phylum of marine invertebrates that have an external bivalved shell to protect their living tissues. With few exceptions, this biomineralized structure is composed of calcite, mixed together with a minor organic fraction, comprising secreted proteins that become occluded in the shell structure, once formed. This organic matrix is thought to display several functions, in particular, to control mineral deposition and to regulate crystallite shapes. Thus, identifying the primary structure of matrix proteins is a prerequisite for generating bioinspired materials with tailored properties. In this study, we employed a proteomic approach to identi…