Project description:A midgut reference sample is compared to midgut samples taken at various stages before (18h, 4h), at (0h), and after (2h, 3h, 4h, 5h, 6h, 8h, 10h, 12h) puparium formation. Keywords = Drosophila, ecdysone, network, genomic, microarray, organogenesis, EcR, midgut, central nervous system, salivary gland, epidermis, imaginal disc, development Keywords: other
Project description:We report the nuclear transcriptome of Drosophila photoreceptor neurons obtained using a tissue-specific approach at different time points after eclosion
Project description:We report the chromatin accessibility landscape of Drosophila photoreceptor neurons obtained using a tissue-specific approach at different time points after eclosion
Project description:Background: Mosquito midgut is an important target for the host parasite interaction studies as it plays a major role in parasite growth and maturation and vector susceptibility. Proteomic approaches coupled with bioinformatics analysis have been used to study expression of functional proteins/enzymes of An. culicifacies susceptible and refractory species midgut in order to understand the mechanism of refractoriness that may help in contributing to unravel the host pathogen interactions. Methodology/Principle findings: In the present study proteomics approaches namely in solution and in gel digestion strategies followed by LC/MS/MS analysis were used. Further bioinformatics analysis were carried out to find out the functional annotated proteins, biological process, molecular function and their sub-cellular location using Gene ontology, SMART analysis, CELLO etc. In solution and in gel approach coupled with LC-MS/MS identified a total of 91 proteins in susceptible species and 69 proteins in refractory species. Comparative analysis between susceptible and refractory species of An. culicifacies indicated that mainly proteins involved in proteolysis mechanism, catalytic activity, peptidases activity and immune related proteins were found to be dominating in refractory as compared to susceptible species. Conclusion/Significance: Based on the present data a significant increase in number of proteins in midgut of refractory An. culicifacies species B were found that may conclude that these proteins may be responsible for the inhibiting parasite growth and linked to the melanization of oocysts or parasite lysis mechanisms in natural populations of refractory mosquito. Hence the progress of our studies at protein level suggests that these identified annotated putative proteins/enzymes may help to explore natural vector-parasite systems and reveal valuable insights into the mechanism of refractoriness which in turn further useful for bringing of novel strategies for control of malaria.
Project description:Drosophila guts were dissociated to single cells as previously described (Dutta et al., 2015) with a few modifications. Guts were dissected from 7-day old adult esg-sfGFP/+, pros- Gal4>RFP/+ females. After pulling out the gut, the crop and midgut/hindgut junction (where the Malpighian tubules branch out of the gut) and Malpighian tubules were removed. Five flies at a time were dissected and immediately transferred into cold PBS containing 1% BSA to avoid exposing the midgut tissue to room temperature for a long period of time. Once 40 guts were dissected, they were transferred to a dissection plate and chopped into small pieces using a razor blade. These small fragments were immediately transferred to an eppendorf tube containing 400 ul of 1 mg/ml elastase/PBS solution (Sigma E0258) and incubated on a shaker at 27 ̊C for 30 min. 1% of BSA in PBS (final concentration) was used to stop the digestion reaction and prevent cells from aggregating. The cell suspension was passed through 100 um and then a 40 um cell strainer and loaded on the top of Optiprep with density gradient 1.12 g/ml. Viable cells were isolated from the top layer of the sample after centrifugation at 800xg for 20 min. Cell viability and number were assessed by 0.4% trypan blue and hemocytometer.
Project description:Obesity has been shown to increase risk for cardiovascular disease and type-2 diabetes. In addition, it has been implicated in aggravation of neurological conditions such as Alzheimer’s. In the model organism Drosophila melanogaster, a physiological state mimicking diet-induced obesity can be induced by subjecting fruit flies to a solid medium disproportionately higher in sugar than protein (HSD) or that has been supplemented with a rich source of saturated fat (HFD). These flies can exhibit increased circulating glucose levels, increased triglyceride content, insulin-like peptide resistance, and behavior indicative of neurological decline, such as decreased climbing ability. We subjected Oregon-R-C flies to variants of the HSD, HFD, or normal (control) diet (ND), followed by a total RNA extraction from fly heads of each diet group for the purpose of Poly-A selected RNA-Sequencing. We targeted at least 50 million paired-end, stranded reads of 75 basepairs in size, and analyzed 4 biological replicates per dietary condition. Our objective was to identify the effects of obesogenic diets on transcriptome patterns, how they differed between obesogenic diets, and identify genes that may relate to pathogenesis accompanying an obesity-like state. Functional annotation and enrichment analysis among genes whose expression was significantly affected by the obesogenic diets indicated an overrepresentation of genes associated with immunity, metabolism, and hemocyanin in the HFD group, and CHK, cell cycle activity, and DNA binding and transcription in the HSD group. Heat map representation of genes affected by both diets illustrated a large fraction of differentially expressed genes between the two diet groups. Diets high in sugar and diets high in fat both have notableeffects on the Drosophila transcriptome in head tissue. The impacted genes, and how they may relate to pathogenesis in the Drosophila obesity-like state, warrant further experimental investigation. Our results also indicate differences in the effects of the HFD and HSD on expression profiles in head tissue of Oregon-R-C flies, despite the reportedly similar phenotypic impacts of the diets.
Project description:Background: Mosquito midgut is an important target for the host parasite interaction studies as this major immune organ plays a role in parasite growth, maturation and vector competence. Most important rural malaria vector of India is Proteomic approaches coupled with bioinformatics analysis have been used to study expression of functional proteins/enzymes of An. culicifacies susceptible and refractory species midgut in order to understand the mechanism of refractoriness that may help in contributing to unravel the host pathogen interactions Methodology/Principle findings: Basic shot gun proteomics approaches along with iTRAQ based differential studies followed by LC/MS/MS were employed to study the proteome of sugar fed An. culicifacies midgut. Further bioinformatics analysis was carried out using Gene ontology, SMART analysis, CELLO algorithm etc. Here proteins identified with iTRAQ Ratio more than 1.5 was considered up regulated and less than 0.67 were considered down regulated. Ratio from 1.5 – 0.67 were considered as moderate to no change. Using these criteria we observed total 80 proteins in An. culicifacies midgut and when compared susceptible species against refractory species, total 30 proteins were found to be downregulated. 3 proteins were found to be upregulated in refractory species Conclusion/Significance: Based on the present data a significant increase in number of proteins in midgut of refractory An. culicifacies species B were found that may conclude that these proteins may be responsible for the inhibiting parasite growth and linked to the melanization of oocysts or parasite lysis mechanisms in natural populations of refractory mosquito. Hence the progress of our studies at protein level suggests that these identified annotated putative proteins/enzymes may help to explore natural vector-parasite systems and reveal valuable insights into the mechanism of refractoriness which in turn further useful for bringing of novel strategies for control of malaria.
Project description:Chronic high sugar feeding induces obesity, hyperglycemia, and insulin resistance in flies and mammals. To gain insight into the mechanisms underlying this response, we profiled gene expression in chronically high sugar fed, wandering (post-prandial) third instar wild type larvae (L3). These data were compared to control-fed larvae as well as those (mid-L3) actively feeding for twelve hours on both diets. We used microarrays to detail the response of Drosophila larvae to high sugar-induced insulin resistance. Male Canton-S third instar larvae were fed control (0.15M) or high (1M) sucrose and selected for RNA extraction and hybridization on Affymetrix microarrays. Wandering L3 were selected as those in the top half of the vial with partial blue guts to confirm that they had stopped eating the (blue) food. Mid-L3 were selected as L2, aged overnight until early L3, then transferred to fresh control or high sucrose food for 12 more hours before selection.