Project description:Curration of small RNAs from four melanogaster-subgroup species (Drosophila simulans, Drosophila sechellia, Drosophila erecta, and Drosophila yakuba) for the purpose of non-coding RNA annotation and comparative genomics assessment.
Project description:The innate immune response of insects relies on several humoral and cellular mechanisms that require the activation of circulating proteases in the hemolymph to be functional. Here, we analyzed the gelatinase and caseinase activities of Drosophila larval hemolymph under normal and pathogenic conditions (bacterial lipopolysaccharides or endoparasitoid Leptopilina boulardi) using in gel zymography. Gelatinase activity was more intense than caseinase activity and qualitative and quantitative variations were observed between D. melanogaster strains and Drosophila species. Mass spectrometry identified a large number of serine proteases in gel bands equivalent to the major gelatinase and caseinase bands and of these, the most abundant and redundant were Tequila and members of the Jonah and Trypsin protease families. However, hemolymph from Tequila null mutant larvae showed no obvious changes in zymographic bands. Nor did we observe any significant changes in hemolymph gelatinases activity 24 h after injection of bacterial lipopolysaccharides or after oviposition by endoparasitoid wasps. These data confirmed that many serine proteases are present in Drosophila larval hemolymph but those with gelatinase and caseinase activity may not change drastically during the immune response.
Project description:Oligoarray analysis was used to determine the number and nature of genes expressed in third-instar Drosophila melanogaster larval midguts. The majority of transcripts were associated with protein synthesis and metabolism. Serine proteases were the main proteolytic enzymes detected. Experiment Overall Design: Three replicates for each sample. We used Affy chip Drosophila 1.
Project description:In Drosophila melanogaster larval hemolymph, under normal conditions, plasmatocytes and crystal cells represent respectively ~95% and ~5% of hemocytes, while lamellocytes, the third larval cell type, are absent since they are only induced after parasitoid wasp oviposition, their role being the encapsulation-melanization response to eliminate the wasp egg. However, even after induction lamellocytes number remains low, making difficult biochemical studies. Here using the D. melanogaster hopTum-l mutant that constitutively produces a high number of hemocytes, we set up a method to purify lamellocytes and analyzed their major proteins by 2D gel electrophoresis and their biotinylated plasma membrane surface proteins by 1D SDS-PAGE after affinity purification. Mass spectrometry allowed to identify 430 proteins from the 2D spots and 344 from affinity purified proteins, totalizing 639 unique proteins. Known lamellocyte markers such as PPO3 and the integrin myospheroid are among the major proteins and affinity purification led to the detection of other integrins and a large array of integrins associated proteins involved in cell-cell junction formation and function. Overall newly identified proteins indicated that these cells are highly adapted to the encapsulation process but may have also several different physiological functions. This study provides the basis for new lamellocyte studies in vivo and in vitro, and develop markers to search whether different populations coexist, establish their origins and decipher their respective roles in drosophila physiology and immunity.
Project description:Bowman-Birk Inhibitor (BBI) has both insecticidal and anti-cancerous properties. It has been hypothesized that dietary BBI slows insect growth by inhibiting the catalytic activity of digestive enzymes trypsins and chyomotrypsins, resulting in the midgut having reduced access to amino acids needed for growth. In mammals, BBI was hypothesized to influence cellular energy metabolism. Thus, we tested the hypothesis that dietary BBI also impacts energy-associated pathways in the midgut of Drosophila melanogaster. We investigated the impact of dietary BBI on the following parameters in the midguts of third-instar Drosophila larvae: (i) cellular metabolites, (ii) global transcriptome response, (iii) putative transcription factor binding sites (TFBSs) associated with the differentially expressed transcripts, and (iv) epithelial cellular structure. Dietary BBI caused: (i) a reduction of cellular DHAP, glucose, and succinate; and, (ii) increased Fructuse-6-phosphate; (ii) differential expression of genes associated with the glucose and fatty acid utilization; and, (iii) a shortening of midgut epithelial microvilli, a phenomenon previously associated with glucose starvation. Additionally, fifty seven percent of the putative TFBSs associated with the differentially expressed transcripts have previously been associated with glucose and insulin activities in mammalian studies. Collectively these results support the hypothesis that dietary BBI influences energy utilization in the Drosophila midgut. Keywords: stress response