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- Binding of insecticidal lectin Colocasia esculenta tuber agglutinin (CEA) to midgut receptors of Bemisia tabaci and Lipaphis erysimi provides cues to its insecticidal potential. [JOURNAL ARTICLE]
- Proteomics 2014 Apr 17.
The insecticidal potential of GNA related lectins against hemipterans has been experimentally proven. However, the basis behind the toxicity of these lectins against hemipterans remains elusive. The present study elucidates the molecular basis behind insecticidal efficacy of Colocasia esculenta tuber agglutinin (CEA) against Bemisia tabaci and Lipaphis erysimi . Confocal microscopic analyses highlighted the binding of 25kDa stable homodimeric lectin to insect midgut. Ligand blots followed by LC MS/MS analyses identified binding partners of CEA as vacuolar ATP synthase and sarcoplasmic endoplasmic reticulum type Ca(2+) ATPase from B.tabaci, and ATP synthase, heat shock protein 70, and clathrin heavy chain assembly protein from L. erysimi. Internalization of CEA into haemolymph was confirmed by western blotting. Glycoprotein nature of the receptors was identified through glycospecific staining. Deglycosylation assay indicated the interaction of CEA with its receptors to be probably glycan mediated. Surface plasmon resonance analysis revealed the interaction kinetics between ATP synthase of B.tabaci with CEA. Pathway prediction study based on Drosophila homologues suggested the interaction of CEA with insect receptors that probably led to disruption of cellular processes causing growth retardation and loss of fecundity of target insects. Thus, the present findings strengthen our current understanding of the entomotoxic potentiality of CEA, which will facilitate its future biotechnological applications. This article is protected by copyright. All rights reserved.
- The fungal chimerolectin MOA inhibits protein and DNA synthesis in NIH/3T3 cells and may induce BAX-mediated apoptosis. [JOURNAL ARTICLE]
- Biochem Biophys Res Commun 2014 Apr 18.
The Marasmius oreades mushroom agglutinin (MOA) is a blood group B-specific lectin carrying an active proteolytic domain. Its enzymatic activity has recently been shown to be critical for toxicity of MOA toward the fungivorous soil nematode Caenorhabditis elegans. Here we present evidence that MOA also induces cytotoxicity in a cellular model system (murine NIH/3T3 cells), by inhibiting protein synthesis, and that cytotoxicity correlates, at least in part, with proteolytic activity. A peptide-array screen identified the apoptosis mediator BAX as a potential proteolytic substrate and further suggests a variety of bacterial and fungal peptides as potential substrates. These findings are in line with the suggestion that MOA and related proteases may play a role for host defense.
- Inductive differentiation of conjunctival goblet cells by γ-secretase inhibitor and construction of recombinant conjunctival epithelium. [JOURNAL ARTICLE]
- Exp Eye Res 2014 Apr 16.
γ-Secretase inhibitor has been shown to promote intestinal goblet cell differentiation. We now demonstrated that the in vitro addition of γ-secretase inhibitor in the culture of human conjunctival epithelial cells significantly promoted the differentiation of conjunctival goblet cells with typical droplet-like phenotype, positive periodic acid-Schiff and goblet cell-specific Muc5Ac, cytokeratin 7 and Helix pomatia agglutinin lectin staining. Moreover, topical application of γ-secretase inhibitor promoted the differentiation of mouse conjunctival goblet cells in vivo. Furthermore, the expression of Notch target gene HES-1 was down-regulated during the differentiation of conjunctival goblet cells. In addition, we found that the recombinant conjunctival epithelium on amniotic membrane showed less goblet cell density and abnormal location when compared with normal conjunctival epithelium, which were improved by the addition of γ-secretase inhibitor in the final induction.
- Fast detection of alpha-fetoprotein-L3 using lens culinaris agglutinin immobilized gold nanoparticles. [Journal Article, Research Support, Non-U.S. Gov't]
- J Nanosci Nanotechnol 2014 Jun; 14(6):4078-81.
Colloidal gold is extensively used for molecular sensing because that the surface plasmon resonance (SPR) bands are affected by changes in the dielectric properties in the close vicinity of these structures due to the binding of ligands to the corresponding receptor molecules immobilized onto the nanostructures through chemi- or physisorption. We describe a simple method for the detection of Alpha-Fetoprotein-L3 which is a new generation of tumor marker for hepatocellular carcinoma (HCC) based on the aggregation of Lens culinaris agglutinin (LCA) Immobilized Gold Nanoparticles. The LCA conjugated GNPs obtained were 15-20 nm in diameter. The visible color change of the gold nanoparticles from purple to blue on interaction with 100 ng/mL of AFP-L3 is the principle applied here for the sensing of AFP-L3 level. UV/Vis spectroscopy also allows assay monitoring by quantifying the red shift of the plasmon resonance wavelength. With this method, the protein AFP-L3 can be rapidly detected as demanded for clinical diagnosis.
- Lectin uptake and incorporation into the calcitic spicule of sea urchin embryos. [JOURNAL ARTICLE]
- Zygote 2014 Apr 15.:1-7.
Summary Primary mesenchyme cells (PMCs) are skeletogenenic cells that produce a calcareous endoskeleton in developing sea urchin larvae. The PMCs fuse to form a cavity in which spicule matrix proteins and calcium are secreted forming the mineralized spicule. In this study, living sea urchin embryos were stained with fluorescently conjugated wheat germ agglutinin, a lectin that preferentially binds to PMCs, and the redistribution of this fluorescent tag was examined during sea urchin development. Initially, fluorescence was associated primarily with the surface of PMCs. Subsequently, the fluorescent label redistributed to intracellular vesicles in the PMCs. As the larval skeleton developed, intracellular granular staining diminished and fluorescence appeared in the spicules. Spicules that were cleaned to remove membranous material associated with the surface exhibited bright fluorescence, which indicated that fluorescently labelled lectin had been incorporated into the spicule matrix. The results provide evidence for a cellular pathway in which material is taken up at the cell surface, sequestered in intracellular vesicles and then incorporated into the developing spicule.
- Decreased core-fucosylation contributes to malignancy in gastric cancer. [Journal Article]
- PLoS One 2014; 9(4):e94536.
The object of the study is to identify N-glycan profiling changes associated with gastric cancer and explore the impact of core-fucosylation on biological behaviors of human gastric cancer cells. A total of 244 subjects including gastric cancer, gastric ulcer and healthy control were recruited. N-glycan profiling from serum and total proteins in gastric tissues was analyzed by DNA sequencer-assisted fluorophore-assisted capillary electrophoresis. The abundance of total core-fucosylated residues and the expression of enzymes involved in core-fucosylation were analyzed with lectin blot, quantitative reverse transcription-polymerase chain reaction, western blot, Immunohistochemical staining and lectin-histochemical staining. The recombinant plasmids of GDP-fucose transporter and α-1,6-fucosyltransferase (Fut8) were constructed and transfected into gastric cancer cell lines BGC-823 and SGC-7901. CCK-8 and wound healing assay were used to assess the functional impact of core-fucosylation modulation on cell proliferation and migration. Characteristic serum N-glycan profiles were found in gastric cancer. Compared with the healthy control, a trianntenary structure abundance, peak 9 (NA3Fb), was increased significantly in gastric cancer, while the total abundance of core-fucosylated residues (sumfuc) was decreased. Core-fucosylated structures, peak6(NA2F) and peak7(NA2FB) were deceased in gastric tumor tissues when compared with that in adjacent non-tumor tissues. Consistently, lens culinaris agglutinin (LCA)-binding proteins were decreased significantly in sera of gastric cancer, and protein level of Fut8 was decreased significantly in gastric tumor tissues compared with that in adjacent non-tumor tissues. Upregulation of GDP-Tr and Fut8 could inhibit proliferation, but had no significant influence on migration of BGC-823 and SGC-7901 cells. Core-fucosylation is down regulated in gastric cancer. Upregulation of core-fucosylation could inhibit proliferation of the human gastric cancer cells.
- CANOMAD responding to weekly treatment with intravenous immunoglobulin (IVIg). [Journal Article]
- BMJ Case Rep 2014.
A 48-year-old man presented with numbness in fingers and diplopia 1 week after a flu-like illness. He made a full recovery but 8 years later developed progressive and disabling sensory ataxia. He had superimposed acute flare-ups with numbness, double vision and ptosis, all following infections. A blood test showed antidisialosyl antibodies including GD1b, GD3, GT1b and GQ1b in keeping with the diagnosis of chronic ataxic neuropathy, ophthalmoplegia, IgM paraprotein, cold agglutinins and antidisialosyl antibodies (CANOMAD). Initial treatment with monthly courses of intravenous immunoglobulin (IVIg) 0.4 g/kg/day for 5 days every 4 weeks helped temporarily but there were marked disabling fluctuations of symptoms. With IVIg 0.6 g/kg/day weekly his symptoms are stable. He remains mobile and has no eye symptoms without need for any other medication. This case demonstrates that weekly IVIg infusions instead of one 5-day course monthly may be able to avoid fluctuations of symptoms in CANOMAD.
- Pathogen bacteria adhesion to skin mucus of fishes. [REVIEW]
- Vet Microbiol 2014 Mar 20.
Fish are always in intimate contact with their environment; therefore they are permanently exposed to very vary external hazards (e.g. aerobic and anaerobic bacteria, viruses, parasites, pollutants). To fight off pathogenic microorganisms, the epidermis and its secretion, the mucus acts as a barrier between the fish and the environment. Fish are surrounded by a continuous layer of mucus which is the first physical, chemical and biological barrier from infection and the first site of interaction between fish's skin cells and pathogens. The mucus composition is very complex and includes numerous antibacterial factors secreted by fish's skin cells, such as immunoglobulins, agglutinins, lectins, lysins and lysozymes. These factors have a very important role to discriminate between pathogenic and commensal microorganisms and to protect fish from invading pathogens. Furthermore, the skin mucus represents an important portal of entry of pathogens since it induces the development of biofilms, and represents a favorable microenvironment for bacteria, the main disease agents for fish. The purpose of this review is to summarize the current knowledge of the interaction between bacteria and fish skin mucus, the adhesion mechanisms of pathogens and the major factors influencing pathogen adhesion to mucus. The better knowledge of the interaction between fish and their environment could inspire other new perspectives to study as well as to exploit the mucus properties for different purposes.
- Central Pupillary Light Reflex Circuits in the Cat: I. The Olivary Pretectal Nucleus. [JOURNAL ARTICLE]
- J Comp Neurol 2014 Apr 5.
The central pathways subserving the feline pupillary light reflex were examined by defining retinal input to the olivary pretectal nucleus (OPt), the midbrain projections of this nucleus, and the premotor neurons within it. Unilateral intravitreal wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) injections revealed differences in the pattern of retinal OPt termination on the two sides. Injections of WGA-HRP into OPt labeled terminals bilaterally in the anteromedian nucleus, and to a lesser extent in the supraoculomotor area, centrally projecting Edinger-Westphal nucleus and nucleus of the posterior commissure. Labeled terminals, as well as retrogradely labeled multipolar cells, were present in the contralateral OPt, indicating a commissural pathway. Injections of WGA-HRP into the anteromedian nucleus labeled fusiform premotor neurons within the OPt, as well as multipolar cells in the nucleus of the posterior commissure. Connections between retinal terminals and the pretectal premotor neurons were characterized by combining vitreous chamber and anteromedian nucleus injections of WGA-HRP in the same animal. Fusiform shaped, retrogradely labeled cells fell within the anterogradely labeled retinal terminal field in OPt. Ultrastructural analysis revealed labeled retinal terminals containing clear spherical vesicles. They contacted labeled pretectal premotor neurons via asymmetric synaptic densities. These results provide an anatomical substrate for the pupillary light reflex in the cat. Pretectal premotor neurons receive direct retinal input via synapses suggestive of an excitatory drive, and project directly to nuclei containing preganglionic motoneurons. These projections are concentrated in the anteromedian nucleus, indicating its involvement in the pupillary light reflex. J. Comp. Neurol., 2014. © 2014 Wiley Periodicals, Inc.
- Identification and IVC of spermatogonial stem cells in prepubertal buffaloes. [JOURNAL ARTICLE]
- Theriogenology 2014 Mar 12.
Development of suitable selective marker for buffalo spermatogonial stem cells (SSCs), optimization of long-term IVC conditions, and their pluripotent retention capacity in buffaloes can be of prime importance in selective genetic modifications of this species. In the present study, we identified CDH1 as a specific marker for buffalo SSCs and revealed that it existed in two protein isoforms (large [135 kDa] and small [90 kDa] subunits) in the buffalo testis; furthermore, immunohistochemical analysis revealed that CDH1 expression was present in spermatogonia but absent in the somatic cells of 4-month-old buffalo testis. After 7 days of enrichment, expression of CDH1 was also detectable in IVC colonies (∼53% enrichment efficiency by Fluorescence-activated cell sorting (FACS)). For long-term culture of SSCs, proliferation studies with different factors showed that combination of 20 ng/mL GDNF, 10 ng/mL FGF2, and 1000 U/mL LIF could significantly promote number of colonies (∼two folds) and proliferation of buffalo SSCs (∼three folds) compared with those of control or single-treatment groups; furthermore, addition of these combination growth factors significantly upregulated the messenger RNA level of spermatogonial-specific and pluripotency-related markers (BCL6B, GFRA1, and POU5F1), whereas downregulated receptor tyrosine kinase (KIT). For confirmation of their stem cell potential, Dolichos biflorus agglutinin-stained cells were identified in the basal membrane of seminiferous tubules of xenotransplanted mice testis. These findings indicate the identification of a new buffalo SSCs marker; furthermore, it may help in establishing long-term culture that would assist in genetic modification of these buffaloes.