Novel biopesticide based on a spider venom peptide shows no adverse effects on honeybees.

Evidence is accumulating that commonly used pesticides are linked to decline of pollinator populations; adverse effects of three neonicotinoids on bees have led to bans on their use across the European Union. Developing insecticides that pose negligible risks to beneficial organisms such as honeybees is desirable and timely. One strategy is to use recombinant fusion proteins containing neuroactive peptides/proteins linked to a 'carrier' protein that confers oral toxicity. Hv1a/GNA (Galanthus nivalis agglutinin), containing an insect-specific spider venom calcium channel blocker (ω-hexatoxin-Hv1a) linked to snowdrop lectin (GNA) as a 'carrier', is an effective oral biopesticide towards various insect pests. Effects of Hv1a/GNA towards a non-target species, Apis mellifera, were assessed through a thorough early-tier risk assessment. Following feeding, honeybees internalized Hv1a/GNA, which reached the brain within 1 h after exposure. However, survival was only slightly affected by ingestion (LD50>100 µg bee(-1)) or injection of fusion protein. Bees fed acute (100 µg bee(-1)) or chronic (0.35 mg ml(-1)) doses of Hv1a/GNA and trained in an olfactory learning task had similar rates of learning and memory to no-pesticide controls. Larvae were unaffected, being able to degrade Hv1a/GNA. These tests suggest that Hv1a/GNA is unlikely to cause detrimental effects on honeybees, indicating that atracotoxins targeting calcium channels are potential alternatives to conventional pesticides.

Glycosylation of beta-1 integrins in B16-F10 mouse melanoma cells as determinant of differential binding and acquisition of biological activity.

Studying B16-F10 cells we could identify beta-1 integrins as laminin, fibronectin and collagen receptors. Gradient ionic strength elution analysis of affinity chromatography showed differential interactions between laminin-binding beta-1 integrins (two beta-1 polypeptides of 105 and 120 kDa) and fibronectin and collagen-binding beta-1 integrins (elution of one major beta-1 polypeptide of 120 kDa) and their respective ligands. To evaluate this diversity we submitted B16-F10 extracts to IEF and SDS-PAGE and found that one beta-1 integrin formed acidic and larger isoforms, while another formed basic and smaller isoforms. To study this difference we also submitted material eluted from WGA-Sepharose columns to IEF but now only the acidic beta-1 isoform was found. Extracts of B16-F10 treated with neuraminidase showed only the basic beta-1 isoform, suggesting that terminal sialic acid residues may be responsible for this acidic pattern, an interpretation supported by the fact that MAA (Maackia ammurensis agglutinin) reacts only with the acidic isoform. Differential glycosylation of beta-1 integrin isoforms in B16-F10 was also demonstrated since the smaller laminin-binding beta-1 integrin isoform reacted only with GNA (Galanthus nivalis agglutinin), whereas the mature larger form reacted with DSA (Datura stramonium agglutinin) and MAA; thus this heterogeneity of beta-1 chains is essentially due to variable glycosylation. Autoradiography and immunoblotting analysis of material separated by 2-dimensional electrophoresis show that only the processed forms of beta-1 integrins are expressed at the cell surface.

A corm-specific gene encodes tarin, a major globulin of taro (Colocasia esculenta L. Schott).

A gene encoding a globulin from a major taro (Colocasia esculenta L. Schott) corm protein family, tarin (G1, ca. 28 kDa) was isolated from a lambda Charon 35 library, using a cDNA derived from a highly abundant corm-specific mRNA, as probe. The gene, named tar1, and the corresponding cDNA were characterized and compared. No introns were found. The major transcription start site was determined by primer extension analysis. The gene has an open reading frame (ORF) of 765 bp, and the deduced amino acid sequence indicated a precursor polypeptide of 255 residues that is post-translationally processed into two subunits of about 12.5 kDa each. The deduced protein is 45% homologous to curculin, a sweet-tasting protein found in the fruit pulp of Curculigo latifolia and 40% homologous to a mannose-binding lectin from Galanthus nivalis. Significant similarity was also found at the nucleic acid sequence level with genes encoding lectins from plant species of the Amaryllidaceae and Lilliaceae families.