Ovalbumin-related Protein X Is a Heparin-binding Ov-Serpin Exhibiting Antimicrobial Activities

Ovalbumin family contains three proteins with high sequence similarity: ovalbumin, ovalbumin-related protein Y (OVAY), and ovalbumin-related protein X (OVAX). Ovalbumin is the major egg white protein with still undefined function, whereas the biological activity of OVAX and OVAY has not yet been explored. Similar to ovalbumin and OVAY, OVAX belongs to the ovalbumin serine protease inhibitor family (ov-serpin). We show that OVAX is specifically expressed by the magnum tissue, which is responsible for egg white formation. OVAX is also the main heparin-binding protein of egg white. This glycoprotein with a predicted reactive site at Lys³⁶⁷-His³⁶⁸ is not able to inhibit trypsin, plasmin, or cathepsin G with or without heparin as a cofactor. Secondary structure of OVAX is similar to that of ovalbumin, but the three-dimensional model of OVAX reveals the presence of a cluster of exposed positive charges, which potentially explains the affinity of this ov-serpin for heparin, as opposed to ovalbumin. Interestingly, OVAX, unlike ovalbumin, displays antibacterial activities against both Listeria monocytogenes and Salmonella enterica sv. Enteritidis. These properties partly involve heparin-binding site(s) of the molecule as the presence of heparin reverses its anti-Salmonella but not its anti-Listeria potential. Altogether, these results suggest that OVAX and ovalbumin, although highly similar in sequence, have peculiar sequential and/or structural features that are likely to impact their respective biological functions.     

Patterned Hydrophobic Domains in the Exopolymer Matrix of Shewanella oneidensis MR-1 Biofilms

Water-dispersible amphiphilic surface-engineered quantum dots (QDs) were found to be strongly accumulated within discrete zones of the exopolymer network of Shewanella oneidensis MR-1 biofilms, but not on the cell surfaces. These microdomains showed a patterned distribution in the exopolymer matrix, which led to a restricted diffusion of the amphiphilic nanoparticles.     

Gap-junction-mediated cell-to-cell communication

Cells of multicellular organisms need to communicate with each other and have evolved various mechanisms for this purpose, the most direct and quickest of which is through channels that directly connect the cytoplasms of adjacent cells. Such intercellular channels span the two plasma membranes and the intercellular space and result from the docking of two hemichannels. These channels are densely packed into plasma-membrane spatial microdomains termed “gap junctions” and allow cells to exchange ions and small molecules directly. A hemichannel is a hexameric torus of junctional proteins around an aqueous pore. Vertebrates express two families of gap-junction proteins: the well-characterized connexins and the more recently discovered pannexins, the latter being related to invertebrate innexins (“invertebrate connexins”). Some gap-junctional hemichannels also appear to mediate cell-extracellular communication. Communicating junctions play crucial roles in the maintenance of homeostasis, morphogenesis, cell differentiation and growth control in metazoans. Gap-junctional channels are not passive conduits, as previously long regarded, but use “gating” mechanisms to open and close the central pore in response to biological stimuli (e.g. a change in the transjunctional voltage). Their permeability is finely tuned by complex mechanisms that have just begun to be identified. Given their ubiquity and diversity, gap junctions play crucial roles in a plethora of functions and their dysfunctions are involved in a wide range of diseases. However, the exact mechanisms involved remain poorly understood.     

Structure prediction of GPCRs using piecewise homologs and application to the human CCR5 chemokine receptor: validation through agonist and antagonist docking

This article describes the construction and validation of a three-dimensional model of the human CC chemokine receptor 5 (CCR5) receptor using multiple homology modeling. A new methodology is presented where we built each secondary structural model of the protein separately from distantly related homologs of known structure. The reliability of our approach for G-protein coupled receptors was assessed through the building of the human C-X-C chemokine receptor type 4 (CXCR4) receptor of known crystal structure. The models are refined using molecular dynamics simulations and energy minimizations using CHARMM, a classical force field for proteins. Finally, docking models of both the natural agonists and the antagonists of the receptors CCR5 and CXCR4 are proposed. This study explores the possible binding process of ligands to the receptor cavity of chemokine receptors at molecular and atomic levels. We proposed few crucial residues in receptors binding to agonist/antagonist for further validation through experimental analysis. In particular, our study provides better understanding of the blockage mechanism of the chemokine receptors CCR5 and CXCR4, and may help the identification of new lead compounds for drug development in HIV infection, inflammatory diseases, and cancer metastasis.     

Temporal variations of metallothionein and metal concentrations in the digestive gland of oysters (Crassostrea gigas) from a clean and a metal-rich site

The concentrations of metallothionein (MT) in bivalves, a potential biomarker of metal pollution, are variable according to specific organs, the highest concentrations being encountered in the digestive glands of oysters. Thus, the present study has been focussed on this organ with a view to validate the use of MT as a biomarker in the field, the temporal changes of metal and metallothionein concentrations have been examined from March to October 1997 in the digestive gland of resident oysters from a clean site (Bay of Bourgneuf, France) and a metal-rich site, the Gironde estuary which has been shown as the most Cd-contaminated marine area in France but is also enriched with Cu and Zn. Moreover, oysters from the clean site have been translocated to the Gironde estuary over the same period. Taking into account all the samples collected over the 7 months of the study, MT concentrations in the digestive gland were positively correlated with weight whereas metal levels were negatively correlated with weight. However, considering monthly samples including specimens from both sites (resident or translocated oysters), a positive correlation was shown between MT and metal concentrations in autumn (September and October) but not in spring and summer. These findings limit the interest of using the digestive gland of oysters as the preferred tissue for the determination of MT concentration as a biomarker.The alternative use of gills should be considered.     

A bit of quiet between the migrations: the resting life of the European eel during their freshwater growth phase in a small stream

The movements and habitat use of resident yellow eels were studied in a stream stretch having both natural and minimum flow zones. N = 12 individuals (total length 505–802 mm) were surgically tagged with radio transmitters and released at their capture sites. They were located using manual radio receivers during the daytime from 2 to 5 days/week over periods ranging from 200 to 329 days, for a total of 1,098 positions. Eels showed home ranges ranging from 33 to 341 m (median value, 62 m), displayed strong fidelity to sites and demonstrated a great degree of plasticity in habitat use. Eels were slightly mobile throughout the year, but their movements were season and temperature dependent, with a maximum during the spring (mean water temperature, 12 °C) and a minimum in winter (3 °C). Stones and roots (utilization rate greater than 50 % of eels for more than 30 % of location days) were significantly the most frequently used habitats. Between the two flow zones, the natural flow was the most occupied, with a significantly higher proportion of resident eels (66.7 % of radio-tagged yellow eels) and longer occupation (81 % of location days) than the minimum flow zone with less suitable habitats.     

Characterization of sphere-propagating cells with stem-like properties from DU145 prostate cancer cells

While accumulating evidence demonstrates the existence of prostate cancer stem cells (PCSCs), PCSCs have not been isolated and thoroughly characterized. We report here the enrichment and characterization of sphere-propagating cells with stem-like properties from DU145 PC cells in a defined serum-free medium (SFM). Approximately 1.25% of monolayer DU145 cells formed spheres in SFM and 26% of sphere cells formed secondary spheres. Spheres are enriched for cells expressing prostate basal and luminal cytokeratins (34βE12 and CK18) and for cancer stem cell markers, including CD44, CD24, and integrin α2β1. Upon culturing spheres under differentiating media conditions in the presence of 10% serum, cells positive for CD44 and CD24 were substantially reduced. Furthermore, spheres could be generated from the sphere-derived adherent cell cultures and xenograft tumors, demonstrating the stemness of DU145 spheres. We have maintained spheres for more than 30 passages within 1.5 years without noticeable loss of their “stemness”. Sphere cells possess self-renewal capacity, display significant increases in proliferation potential, and initiate xenograft tumors with enhanced capacity compared to monolayer DU145 cells. While EGF promoted the generation and maintenance of these stem-like cells, bFGF inhibited these events. Sphere cells proliferate slowly with a significant reduction in the activation of the PI3K-AKT pathway compared to monolayer DU145 cells. While knockdown of PTEN enhanced AKT activation, this did not affect the generation of primary spheres and the propagation of secondary spheres. Consistent with this observation, we were able to demonstrate the generation and propagation of spheres without the addition of external growth factors. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.     

Revisiting iodination sites in thyroglobulin with an organ-oriented shotgun strategy

Thyroglobulin (Tg) is secreted by thyroid epithelial cells. It is essential for thyroid hormonogenesis and iodine storage. Although studied for many years, only indirect and partial surveys of its post-translational modifications were reported. Here, we present a direct proteomic approach, used to study the degree of iodination of mouse Tg without any preliminary purification. A comprehensive coverage of Tg was obtained using a combination of different proteases, MS/MS fragmentation procedures with inclusion lists and a hybrid mass high-resolution LTQ-Orbitrap XL mass spectrometer. Although only 16 iodinated sites are currently known for human Tg, we uncovered 37 iodinated tyrosine residues, most of them being mono- or diiodinated. We report the specific isotopic pattern of thyroxine modification, not recognized as a normal peptide pattern. Four hormonogenic sites were detected. Two donor sites were identified through the detection of a pyruvic acid residue in place of the initial tyrosine. Evidence for polypeptide cleavages sites due to the action of cathepsins and dipeptidyl proteases in the thyroid were also detected. This work shows that semi-quantitation of Tg iodination states is feasible for human biopsies and should be of significant medical interest for further characterization of human thyroid pathologies.     

Peculiarities of <Emphasis Type="Italic">Pycnoporus</Emphasis> species for applications in biotechnology

The genus Pycnoporus forms a cosmopolitan group of four species belonging to the polyporoid white-rot fungi, the most representative group of homobasidiomycetes causing wood decay. Pycnoporus fungi are listed as food- and cosmetic-grade microorganisms and emerged in the early 1990s as a genus whose biochemistry, biodegradation and biotechnological properties have since been progressively detailed. First highlighted for their original metabolic pathways involved in the functionalization of plant cell wall aromatic compounds to yield high-value molecules, e.g. aromas and antioxidants, the Pycnoporus species were later explored for their potential to produce various enzymes of industrial interest, such as hydrolases and oxidases. However, the most noteworthy feature of the genus Pycnoporus is its ability to overproduce high redox potential laccase—a multi-copper extracellular phenoloxidase—as the predominant ligninolytic enzyme. A major potential use of the Pycnoporus fungi is thus to harness their laccases for various applications such as the bioconversion of agricultural by-products and raw plant materials into valuable products, the biopulping and biobleaching of paper pulp and the biodegradation of organopollutants, xenobiotics and industrial contaminants. All the studies performed in the last decade show the genus Pycnoporus to be a strong contender for white biotechnology. In this review, we describe the properties of Pycnoporus fungi in relation to their biotechnological applications and potential.     

Interaction between DMBT1 and galectin 3 is modulated by the structure of the oligosaccharides carried by DMBT1

DMBT1 (deleted in malignant brain tumor 1), a human mucin-like glycoprotein, belonging to the scavenger receptor cystein-rich (SRCR) superfamily, is mainly secreted from mucosal epithelia. It has been shown previously that interaction of hensin, the rabbit ortholog of DMBT1, with galectin 3, a β-galactoside-binding lectin, induces a terminal differentiation of epithelial cells. In this paper, we have used surface plasmon resonance (SPR), to analyse the binding of galectin 3 to two purified samples of human DMBT1:recombinant DMBT1 produced in CHO cells and DMBT1 isolated from intestinal tissues. Characterization of their glycosylation profile by nano-ESI-Q-TOF tandem mass spectrometry showed significant differences in O-glycans between the two DMBT1 samples. Results obtained by SPR demonstrated that the oligosaccharide side chains of DMBT1 are recognized by the carbohydrate-recognition domain (CRD) of galectin 3 and modification in the pattern of oligosaccharides modulates the binding parameters of DMBT1 with galectin 3. Moreover, using immunohistochemistry on paraffin-embedded colonic tissue sections, we could show a co-localisation of DMBT1 and galectin 3 in human intestine, suggesting a potential physiological interaction.