Purpose : The goal of this study was to use RNA Seq to explore the correlation of gene expression of a collection of clinical P. aeruginosa isolates to various phenotypes, such as antimicrobial resistance, biofilm formation or virulence Methods : mRNA profiles were generated for Pseudomonas aerugionsa clinical samples derived from various geographical locations by deep sequencing. The removal of ribosomal RNA was performed using the Ribo-Zero Bacteria Kit (Illumina) and cDNA libraries were generated with the ScriptSeq v2 Kit (Illumina) . The samples were sequenced in single end mode on an Illumina HiSeq 2500 device and mRNA reads were trimmed and mapped to the NC_008463.1 (PA14) reference genome from NCBI using Stampy pipeline with defaut settings. Overall design: mRNA profiles from Pseudomonas aeruginosa derived from liquid LB cultures grown to an OD600 = 2 and deep sequenced using Illumina HiSeq 2500.
Predicting antimicrobial resistance in Pseudomonas aeruginosa with machine learning-enabled molecular diagnostics.
Disease, Subject
View SamplesEstrogen has vascular protective effects in premenopausal women and in women under 60 receiving hormone replacement therapy. However, estrogen also increases risks of breast and uterine cancers and of venous thromboses linked to upregulation of coagulation factors in the liver. In mouse models, the vasoprotective effects of estrogen are mediated by the estrogen receptor alpha (ERa) transcription factor. Here, through next generation sequencing approaches, we show that almost all of the genes regulated by 17-b-estradiol (E2) differ between mouse aorta and mouse liver, and that this is associated with a distinct genomewide distribution of ERa on chromatin. Bioinformatic analysis of E2-regulated promoters and ERa binding site sequences identify several transcription factors that may determine the tissue specificity of ERa binding and E2-regulated genes, including the enrichment of NFkB, AML1 and AP-1 sites in the promoters of E2 downregulated inflammatory genes in aorta but not liver. The possible vascular-specific functions of these factors suggests ways in which the protective effects of estrogen could be promoted in the vasculature without incurring negative effects in other tissues. Our results also highlight the likely importance of rapid signaling of membrane-associated ERa to cellular kinases (altering the activities of transcription factors other than ER itself) in determining tissue specific transcriptional responses to estrogen. Overall design: The aortas or liver fragments of wild-type C57/BL6 mice were incubated ex vivo with 10nM E2 or ethanol vehicle for 4 hours before harvesting for RNA collection. Each condition was performed with two biological replicates, and each replicate contained aortas or liver fragments from 4 mice.
Research resource: Aorta- and liver-specific ERα-binding patterns and gene regulation by estrogen.
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View SamplesThis SuperSeries is composed of the SubSeries listed below.
MicroRNA-Offset RNA Alters Gene Expression and Cell Proliferation.
Specimen part, Treatment
View SamplesMicroRNA-offset RNAs (moRs) were first identified in simple chordates and subsequently in mouse and human cells by deep sequencing of short RNAs. MoRs are derived from sequences located immediately adjacent to microRNAs (miRs) in the primary miR (pri-miR). Currently moRs are considered to be simply a by-product of miR biosynthesis that lack biological activity. Here we show for the first time that a moR is biologically active. We now demonstrate that endogenous and over-expressed moR-21 significantly alters gene expression and inhibits the proliferation of vascular smooth muscle cells (VSMC). We report that the seed region of moR-21 as well as the seed match region in the target gene 3'UTR are indispensable for moR-21-mediated gene down-regulation. We further demonstrated that moR-21-mediated gene repression is Argonaute 2 (Ago2) dependent. In addition, we find that miR-21 and moR-21 may regulate different genes in a given pathway and can oppose each other in regulating certain genes. Taken together, these findings provide the first evidence that microRNA offset RNA regulates gene expression and is biologically active.
MicroRNA-Offset RNA Alters Gene Expression and Cell Proliferation.
Specimen part, Treatment
View SamplesThe genome of vertebrates contains endogenous retroviruses (ERVs) that have resulted from ancestral infections by exogenous retroviruses. ERVs are germline encoded, transmitted in a Mendelian fashion and account for about 8% of the human and 9.9% of the murine genome, respectively1, 2. By spontaneous activation and reintegration ERVs may cause insertional mutagenesis and thus participate in the process of malignant transformation or progression of tumor growth3, 4. However, if the innate immune system is able to recognize and control ERVs has not yet been elucidated. Here we report that, in vitro, nucleic-acid sensing TLRs on dendritic cells are activated by retroviral RNA and DNA from infected cells in vitro. Infection of TLR competent wild type mice with murine leukemia virus (MuLV)-like ERV isolates results in non-canonical gene upregulation, independent of type I IFN. In vivo, TLR3, -7 and -9 triple deficient mice (TLR379-/-) and mice with non functional TLR3, 7 and 9 signaling due to a mutation in UNC93B develop spontaneous ERV-induced viremia. More importantly, in TLR379-/- mice ERV-induced viremia correlates with acute T cell lymphoblastic leukemia (T-ALL). Multiple independent TLR379-/- T cell leukemia lines produce infectious MuLV of endogenous origin. These cell lines display de novo retroviral integration into the Nup214 or Notch1 gene locus leading to gene dysregulation that is reminiscent of aberrant Nup214 and Notch1 expression in human T-ALLs5. Overall, our results demonstrate that in addition to their role in innate immune defense against exogenous pathogens, TLR3,-7, and -9 may be essential for the control of endogenous retroviral mediated T-cell lymphomagenesis.
Nucleic acid-sensing Toll-like receptors are essential for the control of endogenous retrovirus viremia and ERV-induced tumors.
Specimen part
View Samplessubstantial number of people at risk to develop type 2 diabetes could not improve insulin sensitivity by physical training intervention. We studied the mechanisms of this impaired exercise response in 20 middle-aged individuals who performed a controlled eight weeks cycling and walking training at 80 % individual VO2max. Participants identified as non-responders in insulin sensitivity (based on Matsuda index) did not differ in pre-intervention parameters compared to high responders. The failure to increase insulin sensitivity after training correlates with impaired up-regulation of mitochondrial fuel oxidation genes in skeletal muscle, and with the suppression of the upstream regulators PGC1 and AMPK2. The muscle transcriptome of the non-responders is further characterized by an activation of TGF and TGF target genes, which is associated with increases in inflammatory and macrophage markers. TGF1 as inhibitor of mitochondrial regulators and insulin signaling is validated in human skeletal muscle cells. Activated TGF1 signaling down-regulates the abundance of PGC1, AMPK2, mitochondrial transcription factor TFAM, and of mitochondrial enzymes. Thus, increased TGF activity in skeletal muscle can attenuate the improvement of mitochondrial fuel oxidation after training and contribute to the failure to increase insulin sensitivity.
TGF-β Contributes to Impaired Exercise Response by Suppression of Mitochondrial Key Regulators in Skeletal Muscle.
Specimen part
View SamplesThis SuperSeries is composed of the SubSeries listed below.
Matrix elasticity, replicative senescence and DNA methylation patterns of mesenchymal stem cells.
Specimen part, Subject
View SamplesMatrix elasticity influences differentiation of mesenchymal stem cells (MSCs) but it is unclear if these effects are only transient - while the cells reside on the substrate - or if they reflect persistent lineage commitment. In this study, MSCs were continuously culture-expanded in parallel either on polydimethylsiloxane (PDMS) gels of different elasticity or on tissue culture plastic (TCP) to compare impact on replicative senescence, in vitro differentiation, gene expression, and DNA methylation (DNAm) profiles. The maximal number of cumulative population doublings was not affected by matrix elasticity. Differentiation towards adipogenic and osteogenic lineage was increased on soft and rigid biomaterials, respectively - but this propensity was no more evident if cells were transferred to TCP. Global gene expression profiles and DNAm profiles revealed relatively few differences in MSCs cultured on soft or rigid matrices. Furthermore, only moderate DNAm changes were observed upon culture on very soft hydrogels of human platelet lysate. Our results support the notion that matrix elasticity influences cellular differentiation while the cells are organized on the substrate, but it does not have major impact on cell-intrinsic lineage determination, replicative senescence or DNAm patterns.
Matrix elasticity, replicative senescence and DNA methylation patterns of mesenchymal stem cells.
Specimen part, Subject
View SamplesWe used microarrays to detail the global gene expression changes following apical infection of porcine choroid plexus epithelial cells (PCPEC) with Streptococcus suis (S. suis)
In vitro transcriptome analysis of porcine choroid plexus epithelial cells in response to Streptococcus suis: release of pro-inflammatory cytokines and chemokines.
Specimen part
View SamplesGene expression profile was analyzed after knockdown of PAEP in lung cancer cell lines 2106T and H1975 as well as in skin cancer cell line MeWo.
Glycodelin: A New Biomarker with Immunomodulatory Functions in Non-Small Cell Lung Cancer.
Specimen part, Cell line, Treatment
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