Project description:Transcriptional profiling of anterior ovarioles (germaria and round previtellogenic egg chambers) of w1118 virgin females of Drosophila melanogaster 1 to 8 days post eclosion.
Project description:Reactive oxygen species (ROS) serve as intracellular signaling molecules; however, in excess ROS molecules are damaging to biomacromolecules such as DNA, lipids, and proteins. The excess accumulation of ROS leads to oxidative stress, disrupting redox homeostasis in the cell and has been linked with aging and neurodegenerative disease. The eye is particularly vulnerable to oxidative stress due to the tissue’s high energy demand and generation of ROS by products. In proteins, the thiol group on cysteine residues is susceptible to oxidation. Cysteine residues have multiple oxidation states that can be classified into two categories—reversible and irreversible. Irreversible oxidation states include sulfinic and sulfonic acids, which may alter or impair the activity of the protein depending on the position of the cysteine residue. In contrast, reversible oxidation states include sulfenic acid or inter- and intramolecular disulfides, and can often function as redox sensors in the cell. Here, we sought to evaluate how increasing amounts of oxidative stress alters the redox proteome landscape in Drosophila. To characterize the redox proteome, we developed an iodoTMT-multiplex isolation, labeling, and enrichment method to identify cysteine availability and potential oxidation in both S2 cells and w1118 fly eyes. To do this, we exposed S2 cells to 20mM H2O2 relative to control (untreated), and w1118 flies to prolonged blue light versus an untreated control, followed by redox proteome profiling with iodoTMT-sixplex reagents.
Project description:Transcriptional profiling of anterior ovarioles (germaria and round previtellogenic egg chambers) of w1118 virgin females of Drosophila melanogaster 5 to 8 days post eclosion. Anterior ovarioles were dissected from w1118 virgin females 5 to 8 days post eclosion. Three independent biological replicates were prepared for each day. Total RNA was isolated from the samples and used for the generation of multiplexed stranded PolyA+ libraries. External RNA controls were added to ovarian polyA+ RNA during library preparation. 75bp single end libraries were sequenced by Illumina Hiseq 2000.
Project description:Viral infection in Drosophila triggers inducible gene expression, but the function of many of these genes and the biological significance of their induction remain poorly understood. Here, we report the functional characterization of the gene Diedel, which is strongly induced following some viral infections and encodes a 12kDa circulating protein. Diedel mutant flies have reduced viability and succumb rapidly to infection with Sindbis virus. This phenotype is associated with deregulated expression of IMD target genes and is rescued by mutations in the genes imd or IKKg, indicating that Diedel functions as an immunosuppressor cytokine targeting the IMD pathway. Homologues of Diedel are found in the genome of several DNA viruses, providing the first example of virokines in insects. Our results reveal that, besides RNA interference and apoptosis, an NF-kB pathway of inducible defense exerts evolutionary pressure on insect viruses. For each experimental condition (non infected and SINV infected), two biologically independent samples composed of 45 males of w1118 or Diedelcfm null mutant flies were used. Infection has been performed by injecting viral stocks prepared in Tris solution. Flies were then incubated for days at 29°C.
