Vesicle-based secretion in schistosomes: Analysis of protein and microRNA (miRNA) content of exosome-like vesicles derived from Schistosoma mansoni.

Exosomes are small vesicles of endocytic origin, which are released into the extracellular environment and mediate a variety of physiological and pathological conditions. Here we show that Schistosoma mansoni releases exosome-like vesicles in vitro. Vesicles were purified from culture medium by sucrose gradient fractionation and fractions containing vesicles verified by western blot analyses and electron microscopy. Proteomic analyses of exosomal contents unveiled 130 schistosome proteins. Among these proteins are common exosomal markers such as heat shock proteins, energy-generating enzymes, cytoskeletal proteins, and others. In addition, the schistosome extracellular vesicles contain proteins of potential importance for host-parasite interaction, notably peptidases, signaling proteins, cell adhesion proteins (e.g., integrins) and previously described vaccine candidates, including glutathione-S-transferase (GST), tetraspanin (TSP-2) and calpain. S. mansoni exosomes also contain 143 microRNAs (miRNA), of which 25 are present at high levels, including miRNAs detected in sera of infected hosts. Quantitative PCR analysis confirmed the presence of schistosome-derived miRNAs in exosomes purified from infected mouse sera. The results provide evidence of vesicle-mediated secretion in these parasites and suggest that schistosome-derived exosomes could play important roles in host-parasite interactions and could be a useful tool in the development of vaccines and therapeutics.

Extraction of up to 95% of wheat (Triticum aestivum) flour protein using warm sodium dodecyl sulfate (SDS) without reduction or sonication.

Extraction of glutenin polymers without sonication is an essential prerequisite for accurate determination of their composition and molecular size distribution. Sequential fractionation of wheat flour with 0.1 M KCl and 0.25% sodium dodecyl sulfate (SDS) at 21 degrees C and 2% SDS at 60 degrees C extracted up to 95% of total protein. We propose that 2% SDS at 60 degrees C disrupts hydrogen bonds in glutenin and gliadin aggregates, reduces hydrophobic interactions, and facilitates solubilization. Analysis by size-exclusion high-performance liquid chromatography (SE-HPLC), reverse-phase (RP)-HPLC, and SDS-polyacrylamide gel electrophoresis (PAGE) revealed that partitioning of gliadins and glutenins among the extracts differed for two flours with good baking quality (Butte 86 and Jagger) and one with poor baking quality (Chinese Spring). More gliadin was associated with the 0.25% SDS extract for Chinese Spring, whereas more gliadin was associated with the 2% SDS extract for Butte 86 and Jagger. Unextractable glutenin polymer was only 4-5% of total protein for Butte 86 and Chinese Spring and 14% for Jagger.