Project description:We investigate the biological effects of radiation using Drosophila Melanogaster as a model organism, focusing on gene expression and lifespan analysis to determine the effect of different radiation doses. Our results support a threshold effect in response to radiation: no effect on lifespan and no permanent effect on gene expression is seen at doses below 10,000 Roentgens.
Project description:We investigate the biological effects of radiation using Drosophila Melanogaster as a model organism, focusing on gene expression and lifespan analysis to determine the effect of different radiation doses. Our results support a threshold effect in response to radiation: no effect on lifespan and no permanent effect on gene expression is seen at doses below 10,000 Roentgens. Adult male Drosophila were irradiated 2 days after eclosion, with one of 6 radiation doses: 10; 1,000; 5,000; 10,000; 20,000 Roentgens. Samples were taken at 3 time points (2, 10 and 20 days post-irradiation).
Project description:Proteomic response of adult Drosophila melanogaster acclimated at three contrasted thermal conditions (11, 25 and 31 degree Celsius) investigated by 2D-DIGE experiment. Thermal acclimation drastically alters thermotolerance of ectotherms, but the mechanisms determining this plastic response are not fully understood. The present study investigates the proteomic response (2D-DIGE) of adult Drosophila melanogaster acclimated at 11, 25 or 31 °C. As expected 11 °C-acclimation improved cold tolerance and 31 °C-acclimation improved heat tolerance. We hypothesized that the marked organismal responses to acclimation could be detected at the proteomic level assuming that changes in the abundance of specific proteins are linked to the physiological changes underlying the phenotypic response. The 31 °C-acclimated flies displayed a particular divergent proteomic profile where molecular chaperones made up a large number of the proteins that were modulated during heat acclimation. Many other proteins showed significant modulation during acclimation including proteins involved in iron ion and cell redox homeostasis, carbohydrate and energy metabolism, chromatin remodeling and translation, and contractile machinery. Interestingly the changes in protein abundance were often unrelated to transcriptional activity of the genes coding for the proteins, except for the most strongly expressed proteins (e.g. Hsp70). The 11 °C-acclimation evoked weak proteomic response despite the marked effect on the organismal phenotype. Thus the acquired cold tolerance observed here may involve regulatory process such as posttranslational regulation rather than de novo protein synthesis. Bioinformatics and data processing: The proteinScape 2.1 software (BrukerDaltonik GmbH) was used to submit MS/MS data to the following database: NCBI restricted to Drosophila (June 2011, 223,543 sequences) using the Mascot search engine (Mascot server v2.2, http://www.matrixscience.com). Parameters were set as follows: trypsin as enzyme with one allowed miscleavage, carbamidomethylation of cysteins as fixed modification and methionine oxidation as variable modifications. The mass tolerance for parent and fragment ions was set to 0.5 Da. Peptide identifications were accepted if the individual ion Mascot scores were above the identity threshold (the ion score is −10*log(P), where P is the probability that the observed match is a random event, P < 0.05). In case of ambiguous assignments (one compound fit to more than one peptide), peptide were accepted based on the peptide score, meaning that the peptide sequence with the highest score is accepted. The compilation of identified peptides to proteins was performed with the ProteinExtractor algorithm (Thiele et al., 2008, 2010), so that every protein reported was identified by at least one peptide with significant ion Mascot score (above the identity threshold).
Project description:Thermal acclimation study on Drosophila melanogaster reared at 3 different temperatures (12, 25, and 31oC). The proteomic profiles of D. melanogaster under these different temperatures were analyzed and compared using label-free tandem mass spectrometry.
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.