Endogenous oscillator controls surface density of mating receptors in Chlamydomonas eugametos

Summary We describe a circadian rhythm in the surface density of receptors that play a dominant role in the mating process of the unicellular green alga Chlamydomonas eugametos.These receptors — called agglutinins — are large glycoproteins extrinsically bound to the membrane of gamete flagella. We found circadian fluctuations in their density. Since inhibition of protein synthesis affected the agglutinin density without a lag period at any time,we conclude that the density was dependent on de novo synthesis and that the fluctuations in density are caused by circadian oscillations in the rate of agglutinin synthesis. This phenomenon evidently underlies the pronounced endogenous rhythm in mating competence that we described previously (Demets et al. 1987). Finally, we speculate on the nature of the time keeping mechanism that is generating these rhythmic events.     

Humoral antibody response and Ig characterization of the specific agglutinins in rabbits during experimental American trypanosomiasis

A comparative follow up study of the specific agglutinins detected by direct agglutination (DA) test and the immune response detected by specific lysis (SL), indirect immunofluorescence (IFA), indirect hemagglutination (IHA) and complement fixation (CF) tests in rabbits inoculated with trypomastigotes of T. cruzi is reported here.The specific antibody response was detected first by DA test. Reductive cleavage of sera with 2-mercaptoethanol produced a drop in the agglutinin titer of the sera during the first 30 days of infection.The next test to become positive was SL and later on the IFA, IHA and CF tests became positive simultaneously.When fractions obtained by column chromatography in Sephadex G-200 were tested serologically it was demonstrated that specific antibodies were detected mainly in fraction I (IgM) of the pooled rabbit sera obtained 15 days after inoculation (acute stage), and in fraction II (IgG) of the pooled sera obtained from rabbits 90 days after inoculation (chronic stage).Antigens prepared with trypsinized and formolized epimastigotes of three T. cruzi strains, belonging to each one of the different immunological groups described, worked similarly in the detection of specific agglutinin antibodies.Trypanosoma cruzi agglutinins were highly specific in their reaction with their homologous T. cruzi antigens as was proved by the low agglutinin titer obtained in sera from infected rabbits when, instead of T. cruzi epimastigotes, promastigotes of L. donovani were used as antigen, and by the incapacity of this parasite to absorb the T. cruzi agglutinins.     

Genetics of a-agglutunin function in Saccharomyces cerevisiae

The Saccharomyces cerevisiae cell adhesion protein a-agglutinin is composed of an anchorage subunit (Aga1p) and an adhesion subunit (Aga2p). Although functional a-agglutinin is expressed only by a cells, previous results indicated that AGA1 RNA is expressed in both a and cells after pheromone induction. Expression of the Aga2p adhesion subunit in a cells allowed a-agglutinability, indicating that a cells express the a-agglutinin anchorage subunit, although no role for Aga1p in cells has been identified. Most of the a-specific agglutination-defective mutants isolated previously were defective in AGA1; a single mutant (La199) was a candidate for an aga2 mutant. Expression of AGA2 under PGK control allowed secretion of active Aga2p from control strains but did not complement the La199 agglutination defect or allow secretion of Aga2p from La 199, suggesting that the La199 mutation might identify a new gene required for a-agglutinin function. However, the La199 agglutination defect showed tight linkage to aga2::URA3 and did not complement aga2::URA3 in a/a diploids. The aga2 gene cloned from La199 was nonfunctional and contained an ochre mutation. The inability of pPGK-AGA2 to express functional Aga2p in La199 was shown to result from an additional mutation(s) that reduces expression of plasmid-borne genes. AGA2 was mapped to the left arm of chromosome VII approximately 28 cM from the centromere.     

Tyrpanosoma gambiense: phagocytosis in vitro

When Trypanosoma gambiense was exposed in vitro to rat macrophages, the addition of homologous antiserum resulted in immediate adherence of the parasites to the surface of macrophages.The antigenic components of the parasite were fractionated by Sephadex G-200, DEAE-cellulose, and block electrophoresis. Each fraction was used for immunizing rats, and the antisera obtained were each treated for agglutinating antibodies and for their effect on in vitro phagocytosis by rat macrophages. Only the antiserum capable of agglutination could enhance phagocytosis. A strong possibility that the agglutination antigen is mostly responsible for phagocytosis, hence protection, is indicated.Scanning electron micrographs showed that adherence to the macrophage is mostly by the anterior part of the parasite.     

Plant lectins as defense proteins against phytophagous insects

One of the most important direct defense responses in plants against the attack by phytophagous insects is the production of insecticidal peptides or proteins. One particular class of entomotoxic proteins present in many plant species is the group of carbohydrate-binding proteins or lectins. During the last decade a lot of progress was made in the study of a few lectins that are expressed in response to herbivory by phytophagous insects and the insecticidal properties of plant lectins in general. This review gives an overview of lectins with high potential for the use in pest control strategies based on their activity towards pest insects. In addition, potential target sites for lectins inside the insect and the mode of action are discussed. In addition, the effect of plant lectins on non-target organisms such as beneficial insects as well as on human/animal consumers is discussed. It can be concluded that some insecticidal lectins are useful tools that can contribute to the development of integrated pest management strategies with minimal effect(s) on non-target organisms.     

Anti-nutritional effects of a moderate dose of soybean agglutinin in the rat

This study was conducted to investigate the effects of a moderate dose of purified soybean agglutinin on performance and nitrogen digestibility in rats as well as to determine its effects on the protein, DNA and RNA content of the small intestine and pancreas. Twenty-four Sprague - Dawley rats were randomly allotted into one of four groups for a 10-day nitrogen balance experiment. The four groups of rats were fed 7 g of a casein-cornstarch based diet or a similar diet supplemented with 0.1, 0.2 or 0.4 mg/g purified soybean agglutinin. All experimental diets were adjusted to an identical nutrient level. Dose of soybean agglutinin had no significant effect on rat performance. Incorporation of soybean agglutinin in the diet reduced apparent protein digestibility and the utilization of dietary protein by increasing nitrogen loss from the faeces and urine. Fresh pancreatic weight increased in rats fed soybean agglutinin at a level of 0.4 mg/g in the diet compared to the control, but the dry pancreatic weight and the protein content of the pancreas did not differ among the four groups. However the DNA and RNA content of the pancreas had a tendency to increase with a higher level of soybean agglutinin. The weight of the jejunum and its protein, DNA and RNA content were not significantly affected by soybean agglutinin, but the dry weight and the RNA of the jejunum tended to increase with higher levels of soybean agglutinin in the diet. In conclusion, purified soybean agglutinin, at moderate levels in the rats diet, had negative effects on digestive function, such as nitrogen digestibility, nitrogen retention and nitrogen balance. As the level of soybean agglutinin increased, the effects became more pronounced. Meanwhile, hypertrophy of the pancreas was observed with higher doses of soybean agglutinin incorporation in the diets.     

Isolation and composition of the constitutive agglutinins from haploid Saccharomyces cerevisiae cells

Sex-specific agglutinins from the cell surface of haploid cells of Saccharomyces cerevisiae (X2180, mt^a and mt) were purified and analysed. The constitutive agglutinin from mt^a cells was extractable with 3 mM dithiothreitol. It was shown to be a glycoprotein (3% mannose) with an apparent Mr of 43,000 based on gel filtration, but in SDS-PAGE it behaved as a much smaller molecule (Mr between 18,000 and 26,000). About one in three amino acids was a hydroxyamino acid. Its biological activity was resistant to boiling for 1 h, but sensitive to pronase. Intact mt cells retained their agglutinability in the presence of dithiothreitol but limited trypsinizing released a biologically active agglutinin fragment. It had an apparent Mr of 320,000 (gel filtration). When analysed by SDS-PAGE, a single diffuse band with an apparent Mr of 225,000 was observed. The protein was 94% (w/w) mannose with a trace of N-acetyl glucosamine. Its biological activity was almost completely lost after boiling for 1 h. Both agglutinins behaved as monovalent molecules and specifically inhibited the biological activity of both noninduced and pheromone-induced cells. Pheromone treatment of mt^a cells resulted in an apparent 32-fold increase in agglutinin activity at the cell surface, whereas pheromone treatment of mt cells only doubled the apparent agglutinin activity.Abbreviations mt mating type - DTT dithiothreitol - SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis - YPG yeast-peptone-glucose - PAS periodic-acid-Schiff reagent     

Effects of soybean agglutinin on nitrogen metabolism and on characteristics of intestinal tissues and pancreas in rats

Two experiments were conducted to investigate the effects of increasing concentrations of supplemental purified soybean agglutinin on performance, apparent nitrogen digestibility, plasma insulin and cholecystokinine (CCK) levels in rats as well as on the growth of the small intestine and pancreas. In Experiment 1, a 10-day nitrogen balance trial was conducted with 24 male Sprague-Dawley rats (mean BW 85 g) that were randomly allotted to one of four dietary treatments. Rats in each group were provided daily with 7 g of a casein-cornstarch based diet (control) or a diet supplemented with purified soybean agglutinin at 0.4, 0.6 or 0.8 mg/g. Urine and faeces were collected daily and stored at ? 20°C until analysis. In Experiment 2, 30 male Sprague-Dawley rats (mean BW 75 g) were divided into five groups for a 20-day growth experiment. Each rat was fed daily 7 g of a casein-cornstarch based diet (control) or a diet supplemented with purified soybean agglutinin at 0.4, 0.8, 1.2 or 2.0 mg/g. All experimental diets were adjusted to contain a similar level of nutrients. Results from the two experiments showed that supplementation of soybean agglutinin below 2.0 mg/g diet had no significant effect on rat performance. However, rats receiving 2.0 mg soybean agglutinin per gram of diet showed a significant reduction in weight gain compared to the control group. Incorporation of soybean agglutinin in the diet reduced apparent nitrogen digestibility and the retention of dietary nitrogen by increasing nitrogen loss from the faeces and urine. In addition, plasma CCK level increased with increasing inclusion of soybean agglutinin in the diet. On the contrary, the plasma insulin level declined as soybean agglutinin level increased. Soybean agglutinin induced a polyamine-dependent hyperplastic and hypertrophic growth of the small intestine and pancreas by increasing the contents of protein, RNA and DNA, though the increase in weight of small intestine was not significant. Furthermore, 1.2 and 2.0 mg soybean agglutinin per gram of diet promoted proliferation of the jejunum mucosa, while the structure of the brush border epithelium of small intestinal had no damaging change and no diarrhoea was observed in any treatment group. Based on these results, supplementation of low doses of soybean agglutinin or soy protein to parenterally-fed animals affected by atrophic small intestine may promote small intestinal growth.