Sorted wing disc proneural cluster cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Drosophila melanogaster
PROVIDER: GSE2359 | GEO | 2005/03/05
SECONDARY ACCESSION(S): PRJNA91205
REPOSITORIES: GEO
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Project description:Fluorescence-activated cell sorting of M4-GFP wing imaginal disc cells was used to recover a purified population of the cells that comprise the proneural clusters from which sensory organ precursors of the peripheral nervous system (PNS) arise. Whole-genome microarray analysis and in situ hybridization was then used to identify and verify a set of genes that are preferentially expressed in proneural cluster cells. Genes in this set encode proteins with a diverse array of implied functions, and loss-of-function analysis of two candidate genes shows that they are indeed required for normal PNS development.
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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.
2013-12-31 | E-GEOD-41428 | biostudies-arrayexpress
Project description:Genome-wide identification of the binding sites of the Drosophila transcription factors Achaete, Asense, E(spl)m3-HLH and Senseless in wing imaginal cells using DamID profiling.
2014-01-01 | GSE53009 | GEO
Project description:The absent, small or homeotic discs 2 (ash2) gene, a member of the trithorax Group (trxG) of transcriptional regulators, is functionally related to ash1. However, ASH2 and ASH1 belong to distinct multimeric complexes of unknown composition and it is unclear how they act to regulate transcription. In this study, examination of gene expression profiles in wing imaginal discs from ash2 and ash1 mutants revealed their transcriptomes are very similar and correlate with wing phenotypes. Keywords: loss of function analysis
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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.
2013-12-31 | GSE41428 | GEO
Project description:Research on the ability of certain organisms to reconstruct missing structures of their bodies is still in its infancy, despite numerous efforts performed in multiple species. Using expression profile analyses on different timepoints that cover wound healing and regeneration processes (0, 24 and 72 hours post injury), we studied the regenerative behaviour of fragmented wing imaginal discs of D. melanogaster implanted into adult flies. First, through the comparison between cut and intact discs, we identified the effect of implantation on the regeneration process. Filtering this information, we then constructed the specific early regeneration gene signature of wing discs in which multiple transcription factors, immune response genes and members of the JNK, Notch and WNT signaling pathways seem to play an important role. We next compared the transcriptomes of discs 24 and 72 hours after fragmentation to characterize the regenerative machinery and observed a temporal decrease in the cellular metabolic processes, RNA processing and gene expression machinery, suggesting the discs normal activity was stopped on this first interval of time in response to injury and implantation offences. Based on the expression patterns, we elaborated a catalogue of genes involved in wing disc regeneration divided into four classes (wound healing, regeneration, quick response, constant response). Imaginal discs from third instar larvae were 3/4 fragmented, implanted into the abdomen of adult flies and cultured. In order to subtract the noise due to implantation, we repeated the same protocol using intact discs that were cultivated for 24h. Therefore, a total of 12 microarrays were hybridized: four microarrays (with total RNA from non cut wing discs at 0 and 24 hours) were used to assess the effect of the implantation procedure in intact wing discs and the rest (8) were used to measure changes in gene expression in the first 24 hours after disc dissection and implantation (total RNA from wing discs at 0 and 24 hours was hybridized) and during the period between 24 and 72 hours after the cut (total RNA from wing discs at 24 and 72 hours was hybridized). At least two independent total RNA extractions were carried out (using the RNeasy Protect Mini Kit from Qiagen Inc.) for each condition. The four microarrays hybridized for each pair of conditions were done in biological replicate pairs with dyes swapped to take dye bias into account. The whole set of microarrays was normalized following the same protocol, extracting in each case the list of significant genes (at least 1.5-fold change, FDR adjusted pvalue < 0.05).
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Project description:Glioblastoma multiforme (GBM) is an aggressive, heterogeneous and highly vascularized brain tumor. GBM is thought to arise from glioblastoma stem-like cells (GSCs) which are characterized as being either proneural or mesenchymal. The former isolates of GSCs are tight sphere forming and slow growing while mesenchymal GSCs are lose sphere forming, fast growing, highly invasive and when dominant yield poorer patient prognosis. GSCs are known to be plastic in nature and can therefore evolve from a proneural to a mesenchymal state. Here, we observed that factors secreted by endothelial cells (which make up the brain vasculature) alter several properties of GSCs resulting in the acquisition of a more mesenchymal and invasive phenotype coupled with changes at the level of secretory and cellular proteome. Thus, using mass spectrometry, quantitative proteomic analysis and GO term filters, we identified several mesenchymal traits in proneural GSCs exposed to endothelial cell secretome. Specifically, proneural cells treated with the conditioned media derived from human umbilical vein endothelial cells (HUVEC) upregulated the expression of mesenchymal proteins such as CD44 and VIM, while downregulating the expression of the proneural proteins such as NOTCH1, activated NOTCH intracellular domain (NICD), SOX2 and NESTIN, which were validated using flow cytometry (FACS) and western blots (WB). Using DAVID analysis tool, we detected the features of cellular proteome indicative of the activation of NFkB, Wnt and several other pathways in the proneural cells treated with HUVEC conditioned media. Using conditioned media fractionation through several centrifugation steps we identified the extracellular vesicles (EV) sedimented at 100,000 x g using ultracentrifugation, as the source of activity in endothelial conditioned media capable of triggering mesenchymal shift in proneural GSCs. EVs are heterogeneous membrane structures containing multiple bioactive macromolecules, which have the ability to carry multiple bioactive proteins, transfer them to recipient cells and alter their function, signalling and biological programmes. We compared the effects of EVs, soluble fraction and unfractionated conditioned media in terms of their ability to trigger mesenchymal changes in the phenotype of proneural GSCs. Once the cultures were established, the culture medium was removed and replaced with HUVEC-derived material (conditioned media, supernatant or EV fraction) and responses evaluated over 7 days by microscopical analysis of sphere structures, and biochemically by following the aforementioned proneural or mesenchymal markers (WB, FACS). We observed an upregulation of mesenchymal proteins, as well as downregulation of the proneural proteins, mentioned above. These effects recapitulated those of unfractionated conditioned media and were absent from target cells exposed to EV-depleted conditioned media. The data analysis of EV proteome included canonical markers and pathways of cellular vesiculation as well as markers and pathways of interest with regards to the biological effects associated with treatment of GSC recipient cells. In this regard we observed several EV related tetraspanin markers, which were validated using western blot including CD9, CD63, CD81 and a purity control, BIP. Although we identified several potential effectors associated with endothelial cell EVs that could impact proneural cell phenotype, we focused on MMPs for at least three reasons: (i) evidence in the literature (see text) indicated that MMPs may induce differentiation programs in neural stem cells; (ii) MMPs in EV cargo were relatively abundant and have been implicated in various biological processes; (iii) MMPs released from endothelial cells could be functionally involved in disrupting proneural cell sphere structures that we observed in the presence of endothelial cell secretome. We noted the expressions of MMP1, MMP2, MMP11 and MP14 in our HUVEC-EV mass spectrometry dataset, the activity of which was validated using the MMP activity assay kit from abcam (ab112146). Using GO terms we also detected a signal for NFkB pathway activation in the proteome of endothelial (HUVEC) conditioned media-treated proneural GSCs. NFkB activation is regarded as hallmark of mesenchymal phenotype in GSCs and GBM cells. We validated that the upregulation of NFkB, was also true for the proneural cells treated with HUVEC derived EVs. Moreover, upon blocking MMP expression in proneural cells treated with endothelial cell EVs, we inhibited the activation of NFkB activity thereby documenting that the initial effects of MMPs trigger a shift in cellular phenotype toward NfkB activation and mesenchymal reprogramming. Briefly, we compared GO terms of GSC157 cells treated with their own or HUVEC-derived EVs. Validation of the NFkB pathway activation was analysed using WB and immunofluorescence for levels of NFkB and phosphor-NFkB.
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Project description:The distribtion of H3K18ac in the fruit fly genome was analyzed in wing imaginal discs treated for 2 h with etomoxir and compared to control discs.
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