A Large-Scale RNAi-Based Mouse Tumorigenesis Screen Identifies New Lung Cancer Tumor Suppressors that Repress FGFR Signaling
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE47095 | GEO | 2014/05/20
SECONDARY ACCESSION(S): PRJNA203548
REPOSITORIES: GEO
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Project description:We sequenced mRNA from SA cells with SFRS9 knocking down or non-silencing control. Examination of alternative splicing between SFRS9 KD and NS.
2014-05-20 | E-GEOD-47095 | biostudies-arrayexpress
Project description:UnlabelledTo discover new tumor-suppressor genes (TSG), we developed a functional genomics approach in which immortalized but nontumorigenic cells were stably transduced with large-scale shRNA pools and tested for tumor formation in mice. Identification of shRNAs in resulting tumors revealed candidate TSGs, which were validated experimentally and by analyzing expression in human tumor samples. Using this approach, we identified 24 TSGs that were significantly downregulated in human lung squamous cell carcinomas (hLSCC). Amplification of fibroblast growth factor receptor 1 (FGFR1), which aberrantly increases FGFR signaling, is a common genetic alteration in hLSCCs. Remarkably, we found that 17 of the TSGs encode repressors of FGFR signaling. Knockdown of 14 of these TSGs transformed immortalized human bronchial epithelial cells and, in most cases, rendered them sensitive to FGFR inhibitors. Our results indicate that increased FGFR signaling promotes tumorigenesis in many hLSCCs that lack FGFR1 amplification or activating mutations.SignificanceA functional genomics approach identifies new lung TSGs whose loss aberrantly increases FGFR signaling to promote tumorigenesis. These TSGs are frequently downregulated in hLSCCs, indicating that increased FGFR signaling promotes tumorigenesis in many hLSCCs lacking FGFR1 amplification or activating mutations.
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Project description:We sequenced mRNA from SA cells with SFRS9 knocking down or non-silencing control. Examination of alternative splicing between SFRS9 KD and NS.
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Project description:stk25 and tao genes came out of an RNAi screen in the parasitic flatworm Schistosoma mansoni, and these genes shared a similar knockdown phenotype. To test for a functional relationship, we compared transcriptomes of early knockdown worms for each gene, compared to mock controls.
2020-03-11 | GSE146720 | GEO
Project description:During meiosis, the formation of viable haploid gametes from diploid precursors requires that each homologous chromosome pair be properly segregated to produce an exact haploid set of chromosomes. Genetic recombination, which provides a physical connection between homologous chromosomes, is essential in most species for proper homologue segregation. Nevertheless, recombination is repressed specifically in and around the centromeres of chromosomes, apparently because rare centromeric (or pericentromeric) recombination events, when they do occur, can disrupt proper segregation and lead to genetic disabilities, including birth defects. The basis by which centromeric meiotic recombination is repressed has been largely unknown. We report here that, in fission yeast, RNAi functions and Clr4-Rik1 (histone H3 lysine 9 methyltransferase) are required for repression of centromeric recombination. Surprisingly, one mutant derepressed for recombination in the heterochromatic mating-type region during meiosis and several mutants derepressed for centromeric gene expression during mitotic growth are not derepressed for centromeric recombination during meiosis. These results reveal a complex relation between types of repression by heterochromatin. Our results also reveal a previously undemonstrated role for RNAi and heterochromatin in the repression of meiotic centromeric recombination and, potentially, in the prevention of birth defects by maintenance of proper chromosome segregation during meiosis.
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Project description:BackgroundLarge-scale RNAi screening has become an important technology for identifying genes involved in biological processes of interest. However, the quality of large-scale RNAi screening is often deteriorated by off-targets effects. In order to find statistically significant effector genes for pathogen entry, we systematically analyzed entry pathways in human host cells for eight pathogens using image-based kinome-wide siRNA screens with siRNAs from three vendors. We propose a Parallel Mixed Model (PMM) approach that simultaneously analyzes several non-identical screens performed with the same RNAi libraries.ResultsWe show that PMM gains statistical power for hit detection due to parallel screening. PMM allows incorporating siRNA weights that can be assigned according to available information on RNAi quality. Moreover, PMM is able to estimate a sharedness score that can be used to focus follow-up efforts on generic or specific gene regulators. By fitting a PMM model to our data, we found several novel hit genes for most of the pathogens studied.ConclusionsOur results show parallel RNAi screening can improve the results of individual screens. This is currently particularly interesting when large-scale parallel datasets are becoming more and more publicly available. Our comprehensive siRNA dataset provides a public, freely available resource for further statistical and biological analyses in the high-content, high-throughput siRNA screening field.
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Project description:Sulphur is used as a food preservative, especially throughout the table grape and wine industries, <br>however there is increasing concern as to the health rises associated with human consumption. <br>Thus, we investigated the transcript abundance changes in grapes treated with sulfur dioxide and <br>other preservative compounds, compared to control treatments. Export quality, red-skinned M-^QCrimson<br> SeedlessM-^R grapes (Vitis vinifera L.) were harvested at commercial maturity from one vineyard <br>in the Swan Valley region of Western Australia. At least 15 kg grapes per treatment were <br>completely immersed in the treatment solution (? 1 min) and allowed to dry on racks before <br>being weighed in to 7 x 2 kg lined export cartons, without sulphur pads. Once packed, cartons <br>were immediately placed in 2 M-0C storage for up to 56 days and once cool, commercial <br>SO2-generating pads were placed into cartons for SO2 treatment. The salicylic acid (SA,<br> 25 mM), methyl jasmonate (MJ, 5mM) and their combination (25 mM SA + 5 mM MJ) were <br>dissolved in 0.5 % (v/v) dimethyl sulphoxide (DMSO). A 0.5 % solution of DMSO was used<br> as a control treatment. An additional, untreated control for sulphur-treated berries was used.<br>Microarrays were performed in duplicate for the 6 treatments, Sulfur dioxide (plus untreated <br>control), SA, MJ, and the combined SAMJ treatment (plus DMSO control) on grape berries <br>after 21 days of treatment post harvest. The results indcate a large scale re-programing of <br>the grape berry transcritpome, similar to that which has been observed for prolonged <br>exposure to harsh oxidative stress conditions.
2011-03-29 | E-MEXP-3045 | biostudies-arrayexpress
Project description:The ongoing conflict between viruses and their hosts can drive the co-evolution between host immune genes and viral suppressors of immunity. It has been suggested that an evolutionary 'arms race' may occur between rapidly evolving components of the antiviral RNAi pathway of Drosophila and viral genes that antagonize it. We have recently shown that viral protein 1 (VP1) of Drosophila melanogaster Nora virus (DmelNV) suppresses Argonaute-2 (AGO2)-mediated target RNA cleavage (slicer activity) to antagonize antiviral RNAi. Here we show that viral AGO2 antagonists of divergent Nora-like viruses can have host specific activities. We have identified novel Nora-like viruses in wild-caught populations of D. immigrans (DimmNV) and D. subobscura (DsubNV) that are 36% and 26% divergent from DmelNV at the amino acid level. We show that DimmNV and DsubNV VP1 are unable to suppress RNAi in D. melanogaster S2 cells, whereas DmelNV VP1 potently suppresses RNAi in this host species. Moreover, we show that the RNAi suppressor activity of DimmNV VP1 is restricted to its natural host species, D. immigrans. Specifically, we find that DimmNV VP1 interacts with D. immigrans AGO2, but not with D. melanogaster AGO2, and that it suppresses slicer activity in embryo lysates from D. immigrans, but not in lysates from D. melanogaster. This species-specific interaction is reflected in the ability of DimmNV VP1 to enhance RNA production by a recombinant Sindbis virus in a host-specific manner. Our results emphasize the importance of analyzing viral RNAi suppressor activity in the relevant host species. We suggest that rapid co-evolution between RNA viruses and their hosts may result in host species-specific activities of RNAi suppressor proteins, and therefore that viral RNAi suppressors could be host-specificity factors.
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Project description:Large-scale RNAi-based screens are a major technology, but require adequate prioritization and validation of candidate genes from the primary screen. In this work, we performed a large-scale pooled shRNA screen in mouse embryonic stem cells (ESCs) to discover genes associated with oxidative stress resistance and found several candidates. We then developed a bioinformatics pipeline to prioritize these candidates incorporating effect sizes, functional enrichment analysis, interaction networks and gene expression information. To validate candidates, we mixed normal cells with cells expressing the shRNA coupled to a fluorescent protein, which allows control cells to be used as an internal standard, and thus we could detect shRNAs with subtle effects. Although we did not identify genes associated with oxidative stress resistance, as a proof-of-concept of our pipeline we demonstrate a detrimental role of Edd1 silencing in ESC growth. Our methods may be useful for candidate gene prioritization of large-scale RNAi-based screens.
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Project description:BackgroundThe advent of RNA interference techniques enables the selective silencing of biologically interesting genes in an efficient way. In combination with DNA microarray technology this enables researchers to gain insights into signaling pathways by observing downstream effects of individual knock-downs on gene expression. These secondary effects can be used to computationally reverse engineer features of the upstream signaling pathway.ResultsIn this paper we address this challenging problem by extending previous work by Markowetz et al., who proposed a statistical framework to score networks hypotheses in a Bayesian manner. Our extensions go in three directions: First, we introduce a way to omit the data discretization step needed in the original framework via a calculation based on p-values instead. Second, we show how prior assumptions on the network structure can be incorporated into the scoring scheme using regularization techniques. Third and most important, we propose methods to scale up the original approach, which is limited to around 5 genes, to large scale networks.ConclusionComparisons of these methods on artificial data are conducted. Our proposed module network is employed to infer the signaling network between 13 genes in the ER-alpha pathway in human MCF-7 breast cancer cells. Using a bootstrapping approach this reconstruction can be found with good statistical stability. The code for the module network inference method is available in the latest version of the R-package nem, which can be obtained from the Bioconductor homepage.
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