In order to validate our method to develop immortalized cellular models of microglia, we used RNA-seq to confirm the microglial phenotype of a representative clone (C20, a clonal population), which derived from the transformation of human primary microglia (human microglia purchased cryopreserved after purification from Sciencell, Cat. #1900). Transformation was carried out with vesicular stomatitis virus G envelop simian virus 40 large T antigen viral particles (VSVG SV40), containing the pBABE-puro SV40 LT construct (Addgene, Plasmid #13970), followed by VSVG containing the human telomerase reverse transcriptase (hTERT)-neomycin (pBABE-neo-hTERT) construct (Addgene, Plasmid #1774). Immortalized cells were selected in the presence of 2 µg/mL puromycin and 600 µg/mL neomycin. Individual clonal populations, including clone C20, were allowed to grow for approximately 4 weeks prior to further testing. In addition, we also used RNA-seq to verify the capacity of these cellsto respond to an inflammatory stimulus (TNF-a).
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Sex, Specimen part
View SamplesMyotonic dystrophy type 1 (DM1) is an autosomal dominant, CTG microsatellite expansion disease. Transcription of the CTG repeats gives rise to CUG repeat RNA with a toxic gain-of-function. The toxic CUG RNA sequesters the muscleblind-like (MBNL) family of RNA-binding proteins and disrupts their normal cellular function causing global mis-regulation of RNA processing. Multiple approaches have been developed to target the toxic RNA; these include, but are not limited to, displacing MBNL proteins from the CUG repeats, increasing MBNL protein levels or delivery of exogenous MBNL proteins, and blocking the transcription of the CUG repeats. From a screen of diamidine molecules, we previously identified furamidine as a promising lead molecule that was shown to reduce ribonuclear foci and rescue mis-splicing of splicing reporters in a HeLa cell model of DM1. We reported that treatment of the HSALR DM1 mouse model with furamidine partially rescued the Atp2a1 and Clcn1 mis-splicing events via RT-PCR. Here, using RNA-seq examine global splicing, we report that furamidine rescued over 70 mis-splicing events in the HSALR DM1 mouse model and minimally affected gene expression. Heptamidine, in comparison, rescued ~62 events but caused significant alterations in gene expression.
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Sex, Specimen part, Cell line, Treatment
View SamplesWe performed RNA-sequencing on four groups of zebrafish larvae: control, Tg(Myc), Tg(Kras), Tg(Myc)&Tg(Kras) to analyze the expression of genes involved in the lipid-associated pathways.The results revealed high dynamic alterations in almost all aspects of lipid metabolism, among which, the expressions of genes involved in TG/DG/GP transformation and FA desaturation/elongation displayed intensive changes, in consistent with our observations in lipodomics profiling
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View SamplesComparison with antibiotic susceptible and multi-drug resistant Pseudomonas aeruginosa, and responses to antibiotic stresses in multi-drug resistant Pseudomonas aeruginosa
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Specimen part, Cell line
View SamplesMalaria, caused by Plasmodium parasites is responsible for the illness of millions of individuals each year. Plasmodium sporozoites inoculated by mosquitoes migrate to the liver and infect hepatocytes prior to release of merozoites that initiate symptomatic blood-stage malaria. Parasites are thought to be restricted to hepatocytes throughout this obligate liver-stage of replication and differentiation. In contrast to this notion, we found that a subset of hepatic CD11c+ cells co-expressing F4/80, CD103, CD207 and CSF1R, acquired a substantial parasite burden during the liver-stage of malaria, but only after initial hepatocyte infection. These CD11c+ cells found in the infected liver and liver-draining lymph nodes exhibited transcriptionally and phenotypically enhanced antigen-presentation functions; and primed protective CD8 T cell responses against Plasmodium liver-stage restricted antigens. Our findings uncover a novel aspect of Plasmodium biology as well as the fundamental mechanism by which CD8 T cell responses are primed against liver-stage malaria.
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Sex, Specimen part, Cell line, Treatment
View SamplesThe regulatory networks involved in the development of hypoxic and necrotic granulomas following Mycobacterium tuberculosis infections remained elusive. The goal of the study was to determine the role for IL-17 in regulating hypoxic granuloma formation in the lungs of M. tuberculosis-infected C3HeB/FeJ mice.
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Cell line
View SamplesGene expression profile in Calu-3 cells infected with MERS-CoV or SARS-CoV
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View SamplesThe bone marrow-derived macrophages in RPMI1640 containing 10% exo(-) FBS , were treated with apoptotic exosome-like vesicles (10 ug/ml) for 4hr, of which RNA expression was compared with non-treated control cells.
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Sex, Age, Specimen part, Cell line, Treatment
View SamplesXenopus laevis embryos were injected with alpha-amanitin to inhibit RNA polymerase II activity. Embryos were allowed to develop up to stage 10.5 (early gastrula, control and alpha-amanitin injected embryos) and subsequently collected for RNA isolation. The transcriptome profiles of alpha-amanitin injected and control embryos were compared.
Robust activation of a Tbox-Gsc-Otx2 gene network independent of TATA binding protein family members
Compound
View SamplesEpstein-Barr virus (EBV) establishes lifelong infections in its human host. The virus is associated with a broad range of malignancies of lymphoid and epithelial origin, including Burkitt's lymphoma, post-transplant lymphoproliferative disease, nasopharyngeal carcinoma and gastric carcinoma. During the latent phase of its life cycle, EBV expresses more than 40 mature miRNAs that are highly abundant in tumor cells and may contribute to oncogenesis. Although multiple studies have assessed the relative expression profiles of EBV miRNAs in tumor cells, data linking these expression levels to functional target knockdown is mostly lacking. Therefore we set out to systematically assess the EBV miRNA expression levels in EBV+ tumour cell lines, and correlate this to their functional silencing capacity in these cells. We provide comprehensive EBV miRNA expression profiles of the EBV+ cell lines C666-1 (nasopharyngeal carcinoma), SNU-719 (gastric carcinoma), Jijoye (Burkitt's lymphoma), and AKBM (Burkitt's lymphoma) and of EBV- cells ectopically expressing the BART miRNA cluster. By deep sequencing the small RNA population and conducting miRNA-reporter experiments to assay miRNA potency, we were able to compare the expression profiles of the EBV miRNAs with their functional silencing efficacy. We observe a strong correlation between miRNA expression levels and functional miRNA activity. There is large variation in expression levels between EBV miRNAs in a given cell line, whereas the relative expression profiles are well maintained between cell lines. Furthermore, we show that miRNA arm selection bias is less pronounced for viral miRNAs than for human miRNAs. In addition to encoding the largest number of precursor miRNAs of all human herpesviruses, EBV expresses many miRNAs precursors that yield two functional miRNA strands, rather than one guide strand and a non-functional passenger strand. This reduced strand bias may increase the size of the EBV miRNA targetome.
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