An ultraviolet B condition that affects growth and defense in Arabidopsis

Ultraviolet B light (UV-B, 280-315 nm) is the shortest wavelength of the solar spectrum reaching the surface of the Earth. It has profound effects on plants, ranging from growth regulation to severe metabolic changes. Low level UV-B mainly causes photomorphogenic effects while higher levels can induce stress, yet these effects tend to overlap. Here we identified a condition that allows growth reduction without obvious detrimental stress in wild type Arabidopsis rosette plants. This condition was used to study the effects of a daily UV-B dose on plant characteristics of UV-B adapted plants in detail. Exploration of the transcriptome of developing leaves indicated downregulation of genes involved in stomata formation by UV-B, while at the same time genes involved in photoprotective pigment biosynthesis were upregulated. These findings correspond with a decreased stomatal density and increased UV-B absorbing pigments. Gene ontology analysis revealed upregulation of defense related genes and meta-analysis showed substantial overlap of the UV-B regulated transcriptome with transcriptomes of salicylate and jasmonate treated as well as herbivore exposed plants. Feeding experiments showed that caterpillars of Spodoptera littoralis are directly affected by UV-B, while performance of the aphid Myzus persicae is diminished by a plant mediated process

GalNAc/Gal-Binding Rhizoctonia solani Agglutinin Has Antiproliferative Activity in Drosophila melanogaster S2 Cells via MAPK and JAK/STAT Signaling

Rhizoctonia solani agglutinin, further referred to as RSA, is a lectin isolated from the plant pathogenic fungus Rhizoctonia solani. Previously, we reported a high entomotoxic activity of RSA towards the cotton leafworm Spodoptera littoralis. To better understand the mechanism of action of RSA, Drosophila melanogaster Schneider S2 cells were treated with different concentrations of the lectin and FITC-labeled RSA binding was examined using confocal fluorescence microscopy. RSA has antiproliferative activity with a median effect concentration (EC50) of 0.35 µM. In addition, the lectin was typically bound to the cell surface but not internalized. In contrast, the N-acetylglucosamine-binding lectin WGA and the galactose-binding lectin PNA, which were both also inhibitory for S2 cell proliferation, were internalized whereas the mannose-binding lectin GNA did not show any activity on these cells, although it was internalized. Extracted DNA and nuclei from S2 cells treated with RSA were not different from untreated cells, confirming inhibition of proliferation without apoptosis. Pre-incubation of RSA with N-acetylgalactosamine clearly inhibited the antiproliferative activity by RSA in S2 cells, demonstrating the importance of carbohydrate binding. Similarly, the use of MEK and JAK inhibitors reduced the activity of RSA. Finally, RSA affinity chromatography of membrane proteins from S2 cells allowed the identification of several cell surface receptors involved in both signaling transduction pathways

Toxicity and mode of action of fungal lectins in pest insects important in agriculture

The urgent need for more and safer agricultural products, especially food, is rapidly increasing due to an increase of the global human population. Agriculture has suffered from multiple problems, a major threat being insects. These insects have been controlled with different methods mainly by using chemical insecticides. But many problems are associated with the use of these insecticides pushed entomologists to develop alternative methods for these chemical insecticides. Many plant lectins have been reported to possess insecticidal properties but very little is known about the entomotoxic effects of fungal lectins. The main objective of this PhD thesis is to study the insecticidal activity of fungal lectins isolated from two basidiomycetes namely Rhizoctonia solani and Sclerotinia sclerotiorum towards different pest insects and insect cell lines and to investigate the mode of action of these lectins. In vivo assays showed that these lectins have strong insecticidal activity towards pea aphid (Acyrthosiphon pisum) and cotton leafworm (Spodoptera littoralis). While in vitro assays using insect cell lines showed that the activity of these lectins depends on apoptosis induction or inhibition of cell proliferation. These findings suggest that fungal lectins are interesting tool that can be used for bioengineering insect resistance in important agronomical crops

High entomotoxicity and mechanism of the fungal GalNAc/Gal-specific Rhizoctonia solani lectin in pest insects

Whole insect assays where Rhizoctonia solani agglutinin (RSA) was fed to larval stages of the cotton leafworm Spodoptera littoralis and the pea aphid Acyrthosiphon pisum demonstrated a high concentration-dependent entomotoxicity, suggesting that this GalNAc/Gal-specific fungal lectin might be a good control agent for different pest insects. RSA at 10. mg/g in the solid diet of 2nd-instar caterpillars caused 84% weight reduction after 8. days with none of the caterpillars reaching the 4th-instar stage. In sucking aphids, 50% mortality was achieved after 3. days with 9. \u3bcM of RSA in the liquid diet.Feeding of FITC-labeled RSA to both insect pest species revealed strong lectin binding at the apical/luminal side of the midgut epithelium with the brush border zone, suggesting the insect midgut as a primary insecticide target tissue for RSA. This was also confirmed with cell cultures in vitro, where there was high fluorescence binding at the microvillar zone with primary cultures of larval midgut columnar cells of S. littoralis, and also at the surface with the insect midgut CF-203 cell line without lectin uptake in the midgut cells.In vitro assays using insect midgut CF-203 cells, revealed that RSA was highly toxic with an EC50 of 0.3\u3bcM. Preincubation with GalNAc and saponin indicated that this action of RSA was carbohydrate-binding dependent and happened at the surface of the cells. Intoxicated CF-203 cells showed symptoms of apoptosis as nuclear condensation and DNA fragmentation, and this concurred with an increase of caspase-3/7, -8 and -9 activities. Finally, RSA affinity chromatography of membrane extracts of CF-203 cells followed by LC-MS/MS allowed the identification of 5747 unique peptides, among which four putatively glycosylated membrane proteins that are associated with apoptosis induction, namely Fas-associated factor, Apoptosis-linked gene-2, Neuroglian and CG2076, as potential binding targets for RSA. These data are discussed in relation to the physiological effects of RSA