Gene expression regulated by transcription factor MiT in Drosophila
ABSTRACT: To understand the role of MiT in Drosophila, we set out to identify critical gene targets by looking at changes in the WT transcriptome induced by either gain or loss of MiT function. Mutant hindgut and malpighian tubules provided loss-of function tissue and nub-Gal4-driven expression of MiT in the wing epithelium was used for gain-of-function. In the wing disc experiment, 543 genes were upregulated by exogenous MiT, and 359 genes were downregulated (>1.4 fold; P value < 0.01). In the larval HG+MT, 897 genes were downregulated and 898 were upregulated (>1.4 fold; P value < 0.01) after MiT. Among these genes, 85 were both upregulated in wing discs and downregulated in mutant HG+MT, and are the common genes that regulated by MiT in both tissues. We set out to identify potential targets of MiT by comparing the transcriptional profiles of wing discs with and without MiT overexpression (nub-Gal4 UAS-MiT versus nub-Gal4), and separately the profiles of HG+MT tissue from WT and MiT mutant (MiTTZ2/Df(4)TZ). Then, we focused on genetic loci that were upregulated by gain of MiT in the wing and downregulated by loss of MiT in the HG+MT.
Project description:The outcome of Notch proliferation on proliferation depends on the context. In Drosophila wing imaginal discs Notch activation causes hyperplasia despite having localized inhibitory effects on proliferation. To understand te underlying mechanisms we have used genomic strategies to identify the Notch-Su(H) target genes during wing discs hyperplasis. these data are the results from expression profiling the RNAs from hyperplastic wing discs overexpressing Nicd. Direct comparison of third instar lavae wing imaginal disc Nicd (abxUbxFLPase; Act>y>Gal4, UAS GFP; FRT82B tubGal80 with UAS-Nicd; FRT82B) vs control (abxUbxFLPase; Act>y>Gal4, UAS GFP; FRT82B tubGal80 with FRT82B ). 4 Biological replicates, the 2nd replicate was performed as a dye-swap.
Project description:The outcome of Notch activation of proliferation depends on cellular context. In Drosophila wing discs Notch pathway overactivation results in hyperplasia. To understand the mechanisms we have used genomic strategies to indetify the Notch-S(H) target genes directly regulated in wing disc hyperplasia. These data are the results from expression profiling the RNAs from hyperplastic wing discs overexpressing Su(H). Direct comparison of Giant third instar lavae wing imaginal disc (UAS-GFP:Su(H) expressed by the patched[559.1]-Gal4 driver) vs control (UAS-NLS-GFP expressed by the patched[559.1]-Gal4 driver). 3 Biological replicates, the 3rd replicate was performed as a dye-swap.
Project description:The outcome of Notch activation on proliferation depends on cellular context. In Drosophila wing discs Notch activation causes hyperplasia despite having localized inhibitory effects on proliferation. To understand the underlying mechanisms we have used genomic strategies to identify the Notch-Su(H) target genes directly activated during wing disc hyperplasia. These data are the results from ChIP-Chip experiments to identify genomic regions occupied by Su(H) in hyperplastic Nicd-expressing Drosophila wing discs. 2 independent replicates; Immunoprecipitation perfomed with Su(H) antibody on chromatin isolated from wing discs overexpressing Nicd (abxUbxFLPase; Act>y>Gal4, UAS GFP; FRT82B tubGal80 with UAS-Nicd; FRT82B) and compared to the total input DNA.
Project description:We report here the transcriptomic analysis of Drosophila melanogaster wing imaginal discs from third instar female larvae expressing Cyclin G deleted of the PEST domain (the 25 COOH-terminal amin-acids) under the control of the daugterless-Gal4 ubiquitous driver. The negative control was transgenic flies wearing only the daugterless-Gal4 driver. Overall design: Imaginal wing disc mRNA profiles of female UAS-CycGΔP, da-Gal4/+ or da-Gal4/+ Drosophila melanogaster third instar larvae were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.
Project description:Drosophila imaginal disc cells exhibit a remarkable ability to switch cell fates under various perturbations, a phenomenon known as transdetermination (TD). The winged eye (wge) gene induces eye-to-wing TD by its overexpression in eye imaginal discs using eye specific Gal4 driver (eyeless-Gal4). Gene network controlling this process, however, is largely unclear. Additionally, we identified that heterochromatin-related histone methyltransferase Su(var)3-9 is essential for wge-mediated TD. We used microarray to detail the global gene network underlying wge-mediated eye-to-wing TD, and the involvement of Su(var)3-9 in the gene network. Overall design: Third instar larval eye imaginal discs were dissected for RNA extraction and hybridization on Affymetrix microarray. Flies from each genotype were used (I: eyeless-Gal4, II: UAS-wge, III: ey-Gal4; UAS-wge (ey > wge), IV: ey-Gal4 UAS-wge; Su(var)3-91/Su(var)3-92). Three biologic replicates were obtained for each.
Project description:We describe here the genome-wide binding sites of Cyclin G in Drosophila melanogaster wing imaginal discs from third instar female larvae. We used a transgenic line expressing a version Cyclin G deleted of the PEST domain (the 25 COOH-terminal amin-acids) and tagged by Myc under the control of the daugterless-Gal4 ubiquitous driver. Overall design: Genomic binding sites of Myc-CycGΔP in Drosophila melanogaster third instar larvae wing imaginal discs
Project description:The outcome of Notch activation on proliferation depends on cellular context. In Drosophila wing discs Notch activation causes hyperplasia despite having localized inhibitory effects on proliferation. To understand the underlying mechanisms we have used genomic strategies to identify the Notch-Su(H) target genes directly activated during wing disc hyperplasia. These data are the results from ChIP-Chip experiments to identify genomic regions occupied by Su(H) in hyperplastic Su(H)-expressing Drosophila wing discs. 3 independent replicates. Immuno Precipitation perfomed with Su(H) antibody on chromatin isolated from wing discs overexpressing Su(H) (UAS-GFP:Su(H) expressed by the patched[559.1]-Gal4 driver) and compared to the total input DNA. Samples from replicate #1 and #2 were labelled with Cy5 and replicate #3 as dye-swap with Cy3.
Project description:Chromodomains are found in many regulators of chromatin structure. Most of them recognize methylated histones. Here, we investigate the role of the Corto chromodomain. This Drosophila melanogaster Enhancer of Polycomb and Trithorax is involved in both silencing and activation of gene expression. Overexpression of Corto chromodomain (CortoCD) in transgenic flies show that this domain is critical for Corto function and behaves as a chromatin-targeting module. Mass spectrometry analysis of peptides pulled down by CortoCD from nuclear extracts reveals that they correspond to nuclear ribosomal proteins (RPs). Notably, CortoCD binds with high affinity RPL12 tri-methylated on lysine 3 (RPL12K3me3) as demonstrated by real-time interaction analyses. Co-localization of Corto and RPL12 with active epigenetic marks on polytene chromosomes suggests that they are involved in fine-tuning transcription of genes located in open chromatin. Hence, pseudo-ribosomal complexes composed of various RPs might participate in regulation of gene expression in connection with chromatin regulators. RNA-seq analysis of wing imaginal discs overexpressing either Corto or RPL12 show that most deregulated genes are shared by both factors. Interestingly, these common targets are enriched in RP genes suggesting that Corto and RPL12 are involved in dynamic coordination of ribosome biogenesis. To address the role of Corto and RPL12 in regulation of transcription, we deep-sequenced transcripts of wing imaginal discs from third instar larvae over-expressing either FH-cortoCD or RpL12-Myc under control of the wing-specific scalloped::Gal4 driver (sd::Gal4>UAS::FH-cortoCD or sd::Gal4>UAS::RpL12-Myc). Total RNA from FH-cortoCD or RpL12-Myc, the sd::Gal4/+ control or a w1118 reference line were isolated from pools of wing imaginal discs and subjected to RNA-seq on an Illumina high throughput sequencer.
Project description:A 2-hour heat-shock at 37C was used to activate hs-FLP and an actin5C-FRT-stop-FRT-GAL4 transgene in larvae carrying any possible combination of the genetic elements UAS-Myc, UAS-Atu-IR, Max-/-. 48 hours later 16-27 wing imaginal discs were isolated from wandering L3 larvae and polyA-RNA was processed for sequencing.
Project description:Multiple cell types can be specified from a single pool of progenitors through the combinatorial activity of transcriptional regulators, which activate distinct developmental programs to establish different cell fates. The zinc finger transcription factor Glass is required for neuronal progenitors in the Drosophila eye imaginal disc to acquire a photoreceptor identity. Glass is also expressed in non-neuronal cone and pigment cells, but its role in these cells is unknown. To examine how Glass activity is affected by the cellular context, we misexpressed it in different tissues. When expressed in neuroblasts of the larval brain or in epithelial cells of the wing disc, Glass activated both a common core set of target genes and distinct gene sets specific to each tissue. In addition to photoreceptor-specific genes, Glass induced markers of cone and pigment cells. Cell type-specific glass mutations generated in cone or pigment cells using somatic CRISPR revealed autonomous developmental defects, and expressing Glass specifically in these cells partially rescued glass mutant phenotypes. Glass thus acts in both neuronal and non-neuronal cells to promote their differentiation into functional components of the eye, suggesting that it is a determinant of organ identity. Overall design: 6 samples in triplicate, 2 experimental (Glass misexpression (Gl ME) in the wing and Gl ME in the brain), 4 controls (wild-type (WT) eye discs, WT wing discs and WT brains and gl60j brains) Drosophila melanogaster imaginal discs were isolated from Wandering 3rd Instar larvae (W3L) raised at 25C. Gl ME= Glass Misexpression, WT= wild-type. UAS-gl was expressed with tubulin (tub)-GAL4 in clones generated in the wing using Ultrabithorax-Flippase (Ubx-FLP). The inscuteable (insc)-GAL4 driver was used to misexpress Gl in the larval brain in a gl60j background.