Aggregation of sponge cells: 22. Species-specific reactivity of a lectin from the sponge Geodia cydonium

Single cells of the marine sponge Geodia cydonium aggregate species-specifically in the presence of a soluble aggregation factor to form large cell clumps. A lectin isolated from the same sponge species does not cause agglutination of Geodia cells but agglutinates only cells from heterologous species (e.g. Tethya lyncurium, Hemimycale columella, Pellina semitubulosa, Cacospongia scalaris, Verongia aerophoba). The process of agglutination is independent of divalent cations (they do not affect the agglutination process at concentrations up to 50 mM), occurs at 2°C, causes a reduction in the viability of the cells and results in an inhibition of programmed syntheses. The observed differences between the properties of cell agglutination (effect of a lectin in a heterologous system) and cell aggregation (effect of an aggregation factor in the homologous system) is discussed. Cell aggregation is dependent upon the presence of an aggregation factor, the presence of cations and an incubation temperature 2̃0°C; cell aggregation results in a stimulation of programmed syntheses. Cell agglutination requires a heterologous macromolecule (e.g. lectin), it is independent of divalent cations and causes inhibition of programmed syntheses in the cells.     

Further characterization of the sialic acid-binding lectin from the horseshoe crab Carcinoscorpius rotunda cauda

A sialic acid-binding lectin, named carcinoscorpin, has been isolated from the horseshoe crab Carcinoscorpius rotunda cauda. It is a glycoprotein of molecular-weight 420,000, having two subunits of molecular weight 27,000 and 28,000, both subunits responding to glycoprotein stain. Leucine was detected as the only NH₂-terminal amino acid. The sedimentation constant of the native lectin was found to be 12.7 s. On digestion with trypsin, the lectin gave 18 soluble tryptic peptides. This lectin was found to be antigenically unrelated to another sialic acid-binding lectin, limulin, isolated from the horseshoe crab Limulus polyphemus. A lectin-specific disaccharide alcohol namely O-(N-acetylneuraminyl) (2 → 6)2-acetamido-2-deoxy-d-galactitol was found to quench the typical tryptophan fluorescence of the native lectin at 332 nm. The association constant for this interaction was determined spectrofluorimetrically and found to be 1.82 × 10³m^?1.     

Vicia villosa B4 lectin inhibits nucleotide pyrophosphatase activity toward UDP-GalNAc specifically

Plant seed lectins play a defense role against plant-eating animals. Here, GalNAc-specific Vicia villosa B₄ lectin was found to inhibit hydrolysis of UDP-GalNAc by animal nucleotide pyrophosphatases, which are suggested to regulate local levels of nucleotide sugars in cells. Inhibition was marked at low concentrations of UDP-GalNAc, and was reversed largely by the addition of GalNAc to the reaction mixture. In contrast, lectin inhibited enzymatic hydrolysis of other nucleotide sugars, such as UDP-Gal and UDP-GlcNAc, only to a small extent, and GalNAc did not affect such an inhibition. The binding constant of the lectin for UDP-GalNAc was as high as 2.8×10⁵ M^?1 at 4°C, whereas that for GalNAcα-1-phosphate was 1.3×10⁵ M^?1. These findings indicate that lectin inhibition of pyrophosphatase activity toward low concentrations of UDP-GalNAc arises mainly from competition between lectin and enzyme molecules for UDP-GalNAc. This type of inhibition was also observed to a lesser extent with GalNAc-specific Wistaria floribunda lectin, but not apparently with GalNAc-specific soybean or Dolichos biflorus lectin. Thus, V. villosa B₄ lectin shows unique binding specificity for UDP-GalNAc and has the capacity to modulate UDP-GalNAc metabolism in animal cells.     

Schistocephalus solidus and Ligula intestinalis: Pinocytosis by the tegument

Using ruthenium red as a macromolecule, endocytosis was demonstrated in the plerocercoid of Ligula intestinalis and adult Schistocephalus solidus. Uptake, transport across the tegument, and exocytosis across the basal plasma membrane occurred within 6 min. The types of vesicles in the tegument of L. intestinalis are redescribed and their former classification is modified. The vertical and longitudinal distribution of pinosomes in the tegument of adult S. solidus and L. intestinalis plerocercoids were determined. The possible role of macromolecular uptake in the economy of pseudophyllidean tapeworms is discussed with particular reference to growth and defence of an unencysted larval stage in the tissues of the intermediate host.     

Aggregation of sponge cells: a novel mechanism of controlled intercerllular adhesion

From the marine sponge Geodia cydonium a series of macromolecules have been isolated and characterized which are involved in the control of aggregation and separation of sponge cells; these include aggregation factor, aggregation receptor, anti-aggregation receptor, β-glucuronidase, β-glucuronosyltransferase, β-galactosyltransferase, β-galactosidase and a lectin. These components might be linked in the following sequence: (a) activation of the aggregation receptor by its enzymic glucuronylaion; (b) adhesive recognition of the cells, mediated by the aggregation factor and the glucuronylated aggregation receptor; (c) inactivation of the aggregation receptor by its deglucuronylation with the membrane-associated β-glucuronidase; (d) cell separation due either to the loss of the recognition site (glucuronic acid) of the aggregation receptor for the aggregation factor or to an inactivation of the aggregation factor by the anti-aggregation receptor. The activity of the anti-aggregation receptor is probably controlled by the Geodia lectin.     

A phytohaemagglutinin from the Azolla-anabaena symbiosis

Proteins recovered from cell-free extracts of the Azolla-Anabaena azollae symbiosis exhibited haemagglutination activity; galactose was the most effective carbohydrate tested in preventing haemagglutination. Extracts of cyanobacteria-free Azolla also caused haemagglutination but extracts of free-living or symbiotic Anabaena azollae did not. Symbiotic Azolla plants grown on NO₃^? showed lower haemagglutination activity than did those grown on N₂; activity increased on removal of NO₃^?. The lower activity of the NO₃^?-grown material may be due to NO₃su? exerting a direct effect on lectin activity/synthesis, or it may act indirectly by inhibiting the development of Anabaena which in turn affects the Azolla lectin. The purified lectin was shown to be composed of 6 sub-units, each of M.W. 21000.     

Nudibranch-sponge feeding dynamics: Benefits of symbiont-containing sponge to Archidoris montereyensis (Cooper, 1862) and recovery of nudibranch feeding scars by Halichondria panicea (Pallas, 1766)

Selected interactions between the encrusting sponge Halichondria panicea and its primary predator, the dorid nudibranch Archidoris montereyensis, were investigated in a high-latitude rocky intertidal community spatially dominated by H. panicea. Feeding experiments were conducted in which A. montereyensis pairs were provided with sponge containing symbiotic zoochlorellae or sponge in which the zoochlorellae population had been reduced or removed by shading. Nudibranchs consuming H. panicea with symbiotic zoochlorellae had higher feeding, growth, and egg production rates than individuals eating aposymbiotic sponge. We simulated A. montereyensis predation on H. panicea by creating typically sized feeding grooves in the sponge. H. panicea's response was high linear growth rates into the experimental feeding grooves, generally recovering most of the groove area within 4 weeks. Overall, the sponge's rapid response to tissue damage minimizes grazing impacts and substrate loss and reduces susceptibility to wave removal.     

Arabinogalactans of sexual and somatic tissues of gladiolus and lilium

Arabinogalactans are present in extracts of the sexual and somatic tissues of two monocotyledons, Gladiolus and Lilium. The arabinogalactans precipitated from plant extracts by the β-glucosyl artificial antigen (Yariv antigen) show differences in the proportions of the major monosaccharides, galactose and arabinose. Gladiolus stigma and style preparations contain a single arabinogalactan component, but the other tissues examined contained at least two distinct groups of arabinogalactans. These groups can be separated electrophoretically or by lectin affinity chromatography.     

Studies on the relationship between lectins from Axinella polypoides agglutinating bacteria and human erythrocytes

A new lectin, called Axinella IV, with bacteria agglutinating activity was detected in the crude extract of the sponge Axinella polypoides. Immunofluorescence studies and immunoelectrophoresis revealed no cross-reactivities with the already known lectins Axinella I. II, and III. d-Galactose and oligosaccharides with d-Galactose in terminal nonreducing position are inactive in agglutination inhibition tests demonstrating also different specificity of Axinella IV lectin when compared with the three aforementioned lectins.     

Sponge predation in the oyster reef community as demonstrated with Cliona celata Grant

Predation by various invertebrates on the burrowing sponge, Cliona celata Grant, was investigated both in the laboratory and field, in order to determine the importance of predation to a sponge population and the adaptive advantage of burrowing to escape predators. Thirty-one species of molluscs, crustaceans, polychaetes, and echinoderms commonly found on oyster beds in Beaufort, North Carolina were tested for their ability to prey on C. celata. Gastropods Diodora cayenensis Lamarck, Seila adamsi, H. C. Lea and Doriopsilla pharpa Marcus, isopod Cilicaea caudata (Say), decapods Alpheus heterochaelis Say, Panopeus herbsti Milne Edwards, Neopancpe sayi (Smith) Eurypanopeus depressus (Smith), Menippe mercenaria (Say) and Pilutnnus sayi Rathbun, and echinoids Arbacia punctulata (Lamarck) and Lytechinus variegatus (Lamarck) were all found to ingest sponge. Diodora, Seila, Doriopsilla, Cilicaea, Alpheus and Arbacia frequently eat sponges in nature. All sponge predators were able to obtain sponge ventilation papillae, which extend beyond the surface of the shell housing the sponge. Papillae lost to predation were regenerated within 12 days in Cliona celata; little regeneration was noted in predator-damaged Haliclona permollis (Bowerbank), a nonburrowing sponge. Only Arbacia was able to breach sponge galleries in the shell and destroy Cliona celata 2 to 3 times faster than growth could replace lost sponge tissue. The paucity of subtidal shelly bottom sponge fauna and cryptic or high intertidal habits of oyster bed sponges in Beaufort Harbor suggest at least partial control of populations of several common sponges by predators.     

Isolation and characterization of a lectin from the seeds of Erythrina Edulis

A galactose-specific lectin was isolated from the seeds of Erythrina edulis. The protein was purified by affinity chromatography of the globulin fraction on an allyl-galactoside polyacrylamide gel. The hemagglutination properties, amino acid composition, A₂₈₀, MW of the protein and of its subunits, carbohydrate content, electrophoretic pattern and isoelectric point were determined. Comparison of its properties with those of other Erythrina lectins shows that the protein is a distinct member of this group of lectins.     

Bitten down to size: Fish predation determines growth form of the Caribbean coral reef sponge Mycale laevis

Interactions between organisms add complexity to ecosystem function, particularly on coral reefs. The Caribbean orange icing sponge Mycale laevis is semi-cryptic, often growing under coral colonies or between coral branches. This association is reportedly a mutualism, with the sponge deterring boring sponges from invading the coral skeleton and the coral providing an expanding surface for sponge growth. But is there an alternative explanation for the proximity of sponge and coral? We examined the importance of fish predation on the growth of the sponge. While the semi-cryptic growth form of M. laevis predominates on reefs off the Florida Keys and the Bahamas Islands, M. laevis grows with a non-cryptic, erect morphology off Bocas del Toro, Panama. Surveys revealed that sponge-eating fishes were rare or absent at Bocas del Toro compared to sites in the Florida Keys. Because past studies were inconsistent about the palatability of M. laevis to fish predators, we conducted feeding experiments with sponges from all three sites. Crude organic extracts of M. laevis from all three sites were palatable to generalist fish predators in aquarium assays, and field feeding assays and caging experiments conducted in the Florida Keys confirmed that spongivorous fishes readily ate exposed fragments of M. laevis. Our results suggest that M. laevis is restricted to its semi-cryptic growth form by spongivorous predators, with corals providing a physical refuge from predation. This alternative explanation supports the broader hypothesis that Caribbean reef sponges can be categorized on the basis of chemical defense into defended, palatable, and preferred species, the last of which are restricted to refugia.     

Interaction between Dolichos biflorus lectin and chick embryonic fibroblasts at different stages of development

The interaction between chick embryo fibroblasts and A1-specific blood group Dolichos biflorus lectin has been studied at various stages of embryo development. The site number (($\text{0.26±0.03) · 10}{}^6\text{sites/cell}$) remains the same during development whereas the affinity constant apparently decreases from 8-day cells onwards. The effects of cell number, temperature and time course on the Dolichos binding to fibroblasts were not age dependent. Competitive binding experiments revealed that Dolichos receptor sites were distinct from binding sites of Robina pseudoacacia lectin and concanavalin A, but partially related to binding sites of Ricinus lectin. Thymidine incorporation by fibroblasts in the presence of Dolichos lectin was age dependent. It was inhibited in 6-day cells and weakly stimulated in 16-day cells, but not modified in 12-day cells. Dolichos lectin effects on embryo fibroblasts were very specific because both binding to cells and effect on thymidine incorporation were blocked by $\text{N-}\text{acetylgalactosamine}$, the determinant of Dolichos lectin, as well as by Dolichos antiserum.     

A new Bauhinia monandra galactose-specific lectin purified in milligram quantities from secondary roots with antifungal and termiticidal activities

This work describes the purification in milligram quantities of a lectin from Bauhinia monandra secondary roots (BmoRoL) and its antifungal and termiticidal activities. The BmoRoL (6.2 mg) was isolated through ammonium sulfate fractionation and affinity chromatography on guar gel. Native lectin was resolved as a single band on polyacrylamide gel electrophoresis for basic proteins. Under denaturing and reducing conditions it appeared as a unique glycosylated polypeptide of 26 kDa. The highest agglutination activity of BmoRoL was found with glutaraldehyde-treated rabbit erythrocytes. BmoRoL showed antifungal activity against phytopathogenic species of Fusarium and was more active on Fusarium solani. The lectin also showed termiticidal activity on Nasutitermes corniger workers and soldiers with LC₅₀ of 0.09 and 0.395 mg ml⁻¹ for 12 days. In conclusion, BmoRoL is a new antifungal and termiticidal lectin that can be purified in milligram quantities and has potential biotechnological application for control of agricultural pests.     

Structural characterization of a galectin isolated from the marine sponge Chondrilla caribensis with leishmanicidal potential

BackgroundSolving primary structure of lectins leads to an understanding of the physiological roles within an organism and its biotechnological potential. Only eight sponge lectins have had their primary structure fully determined.MethodsThe primary structure of CCL, Chondrilla caribensis lectin, was determined by tandem mass spectrometry. The three-dimensional structure was predicted and the protein-carbohydrate interaction analysed by molecular docking. Furthermore, the anti-leishmanial activity was observed by assays with Leishmania infantum.ResultsThe amino acid sequence consists of 142 amino acids with a calculated molecular mass of 15,443 Da. The lectin has a galectin-like domain architecture. As observed in other sponge galectins, the signature sequence of a highly conserved domain was also identified in CCL with some modifications. CCL exhibits a typical galectin structure consisting of a β-sandwich. Molecular docking showed that the amino acids interacting with CCL ligands at the monosaccharide binding site are mostly the same as those conserved in this family of lectins. Through its interaction with L. infantum glycans, CCL was able to inhibit the development of this parasite. CCL also induced apoptosis after eliciting ROS production and altering the membrane integrity of Leishmania infantum promastigote.ConclusionsCCL joins the restricted group of sponge lectins with determined primary structure and very high biotechnological potential owing to its promising results against pathogens that cause Leishmaniasis.General significanceAs the determination of primary structure is important for biological studies, now CCL can become a sponge galectin with an exciting future in the field of human health.     

Monovalent and bivalent N-fucosyl amides as high affinity ligands for Pseudomonas aeruginosa PA-IIL lectin

The adhesion of bacteria to human glycoconjugates can be inhibited by soluble glycomimetics that compete with the natural target. Four monovalent and one divalent α-fucosyl amides have been tested for their affinity for a fucose-binding lectin from Pseudomonas aeruginosa. Isothermal calorimetric titrations demonstrated that they bind to the lectin in the micromolar range, with highest affinity for the divalent ligand. Molecular modelling established that, compared to Ο-fucoside compounds, the glycomimetic amide group resulted in the loss of water-bridged hydrogen bonds that could be partially compensated by additional contact of the aglycone with the protein surface.     

Interaction of pneumococcal S-14 polysaccharide with lectins from Ricinus communis,Triticum vulgaris, and bandeiraea simplicifolia

Two purified lectins, namely, wheat-germ agglutinin (from Triticum vulgaris) and the hemagglutinin from Ricinus communis seeds, readily form a precipitate with pneumococcal S-14 polysaccharide, whereas the Bandeiraea simplicifolia lectin (BS 1) does not. Exhaustive periodate oxidation and borohydride reduction of S 14 modifies terminal β-D-galactopyranosyl residues, as well as chain D-glucopyranosyl residues, and abolishes reactivity with both the R. communis lectin and wheat-germ agglutinin. Controlled periodate oxidation followed by Smith degradation cleaves only terminal β-D-galactopyranosyl residues, giving a linear polymer, the structure of which was determined by methylation analysis. This derived polymer, containing (1→6)-linked 2-acetamido-2-deoxy-β-D-glucosyl residues, readily precipitated wheat-germ agglutinin, but not the R. communis lectin.     

Mapping glycoconjugate-mediated interactions of marine Bacteroidetes with diatoms

The degradation of diatoms is mainly catalyzed by Bacteroidetes and this process is of global relevance for the carbon cycle. In this study, a combination of catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) and fluorescent lectin binding analysis (FLBA) was used to identify and map glycoconjugates involved in the specific interactions of Bacteroidetes and diatoms, as well as detritus, at the coastal marine site Helgoland Roads (German Bight, North Sea). The study probed both the presence of lectin-specific extracellular polymeric substances (EPS) of Bacteroidetes for cell attachment and that of glycoconjugates on diatoms with respect to binding sites for Bacteroidetes. Members of the clades Polaribacter and Ulvibacter were shown to form microcolonies within aggregates for which FLBA indicated the presence of galactose containing slime. Polaribacter spp. was shown to bind specifically to the setae of the abundant diatom Chaetoceros spp., and the setae were stained with fucose-specific lectins. In contrast, Ulvibacter spp. attached to diatoms of the genus Asterionella which bound, among others, the mannose-specific lectin PSA. The newly developed CARD-FISH/FLBA protocol was limited to the glycoconjugates that persisted after the initial CARD-FISH procedure. The differential attachment of bacteroidetal clades to diatoms and their discrete staining by FLBA provided evidence for the essential role that formation and recognition of glycoconjugates play in the interaction of bacteria with phytoplankton.     

Evolutionary divergence of the paralogs Methoprene tolerant (Met) and germ cell expressed (gce) within the genus Drosophila

Juvenile hormone (JH) signaling underpins both regulatory and developmental pathways in insects. However, the JH receptor is poorly understood. Methoprene tolerant (Met) and germ cell expressed (gce) have been implicated in JH signaling in Drosophila. We investigated the evolution of Met and gce across 12 Drosophila species and found that these paralogs are conserved across at least 63 million years of dipteran evolution. Distinct patterns of selection found using estimates of dN/dS ratios across Drosophila Met and gce coding sequences, along with their incongruent temporal expression profiles in embryonic Drosophila melanogaster, illustrate avenues through which these genes have diverged within the Diptera. Additionally, we demonstrate that the annotated gene CG15032 is the 5′ terminus of gce.In mosquitoes and beetles, a single Met-like homolog displays structural similarity to both Met and gce, and the intron locations are conserved with those of gce. We found that Tribolium and mosquito Met orthologs are assembled from Met- and gce-specific domains in a modular fashion. Our results suggest that Drosophila Met and gce experienced divergent evolutionary pressures following the duplication of an ancestral gce-like gene found in less derived holometabolous insects.     

Lectin receptors in Trypanosoma cruzi an N-acetyl-d-glucosamine-containing surface glycoprotein specific for the trypomastigote stage

We have investigated the interaction of three lectins, differing in their sugar specificities, with the surface of the three differentiation stages of Trypanosoma cruzi. The Scatchard constants for each lectin and parasite stage imply that differentiation of T. cruzi is accompanied by changes in the cell surface saccharides. Trypomastigotes obtained from two different sources do not differ appreciably as to the number and affinity of binding sites for the three lectins employed, suggesting a similar cell-surface saccharide composition. These conclusions are reinforced by sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of the ¹³¹I-labeled surface glycoproteins, following isolation by affinity chromatography. The surface membrane of trypomastigotes, the infective stage to T. cruzi for mammalian cells, possesses a specific glycoprotein of apparent M_r 85 000 (Tc-85) which is absent from the other two stages and can be isolated by affinity chromatography on wheat germ agglutinin-Sepharose columns. This glycoprotein also binds to concanavalin A, but not to Lens culinaris lectin. The binding of Tc-85 to wheat germ agglutinin is unnafected by treatment of either the isolated glycoprotein or intact living trypomastigotes with neuraminidase. Since $\text{N-}\text{acetyl}\text{-}\text{d}\text{-}\text{glucosamine}$ inhibits internalization of trypomastigotes by cultured mammalian cells, it is suggested that Tc-85 might be involved in adhesion and/or interiorization of T. cruzi into mammalian cells, possibly via recognition of an ubiquitous host-cell surface $\text{N-}\text{acetyl}\text{-}\text{d}\text{-}\text{glucosamine}\text{-}\text{specific}$ receptor activity.