Transcript dynamics in mitotic exit mutants in the S. cerevisiae BF264-15D strain background. We examined the extent to which periodic cell-cycle transcription persisted in cells arrested in anaphase with intermediate level of B-cyclins.
Reconciling conflicting models for global control of cell-cycle transcription.
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View SamplesIn this study we examined the influence of seminal plasma on gene expression in human Ect1 ectocervical epithelial cells, and the extent to which recombinant TGF3 elicits comparable changes. Ect1 cells were incubated with recombinant human TGF3 (5 ng/ml), 10% pooled human seminal plasma (v/v), or medium alone for 10h. RNA was reverse transcribed into cDNA and hybridized to Affymetrix GeneChip Human Genome U133 plus 2.0 microarrays (Affymetrix, Santa Clara, CA). Exposure of Ect1 cells to seminal plasma resulted in differential expression of a total of 3955 probe sets, identified using high stringency criteria with MAS 5.0 analysis. These corresponded to 1338 genes up-regulated and 1343 genes down-regulated by seminal plasma. TGF3 treatment of Ect1 cells resulted in differential expression of 884 probe sets, corresponding to 346 up-regulated genes and 229 down-regulated genes. The genes differentially regulated by seminal plasma included several genes associated with cytokinecytokine receptor interaction, TGF signalling, JAK/STAT signalling or VEGF signalling pathways, as specified by the KEGG database. Of 47 genes in these families, 17 (36.1%) were similarly regulated by both seminal plasma and TGF3. These data, together with additional experiments showing all three TGF isoforms can regulate inflammatory cytokine expression in Ect1 cells, identify TGF isoforms as key agents in seminal plasma that signal induction of pro-inflammatory cytokine synthesis in cervical cells.
TGF-β mediates proinflammatory seminal fluid signaling in human cervical epithelial cells.
Cell line, Treatment
View SamplesWe used morpholinos and gene editing tools to inactivate the srsf5a gene. In contrast to srsf5a homozygous mutants that did not display any phenotypic traits, microinjection of sMOsrsf5a led to developmental defects. By using RNA sequencing on morphants and control embryos we were able to identify a plethora of morpholino inadvertant target. Overall design: Two biological replicates were used per conditions. Samples named CtrlMO consist in embryos injected with the control morpholino (5''-CCTCTTACCTCAGTTACAATTTATA-3'', Gene Tools). Samples named sMOsrsf5a consist in embryos injected with the morpholino against srsf5a (5''-GGATTCAGTCTCACCTCTCACTGCA-3'', Gene Tools).
Number of inadvertent RNA targets for morpholino knockdown in Danio rerio is largely underestimated: evidence from the study of Ser/Arg-rich splicing factors.
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View SamplesAbstract of publicaton: CD4/CD8 double-positive (DP) thymocytes express the transcriptional repressor Histone Deacetylase 7 (HDAC7), a class IIa HDAC that is exported from the cell nucleus after T cell receptor (TCR) engagement. Through signal-dependent nuclear export, class IIa HDACs such as HDAC7 mediate signal-dependent changes in gene expression that are important to developmental fate decisions in multiple tissues. We report that HDAC7 is exported from the cell nucleus during positive selection in thymocytes, and regulates genes mediating the coupling between TCR engagement and downstream events that determine cell survival. Thymocytes lacking HDAC7 are inefficiently positively selected due to a severely shortened lifespan and exhibit a truncated repertoire of TCR Jalpha segments. The expression of multiple important mediators and modulators of the response to TCR engagement is altered in HDAC7-deficient thymocytes, resulting in increased tonic MAP kinase activity that contributes to the observed loss of viability. Remarkably, the activity of Protein Kinase D, the kinase that mediates nuclear export of HDAC7 in response to TCR signaling, is also increased in HDAC7-deficient thymocytes, suggesting that HDAC7 nuclear export governs a self-sustaining auto-excitatory loop. These experiments add to the understanding of the life/death decision in thymic T cell development, define a novel function for class IIa HDACs, and point to a novel feed-forward mechanism whereby these molecules regulate their own state and mediate stable developmental transitions. Title of manuscript: Nuclear Export of Histone Deacetylase 7 During Thymic Selection Mediates Immune Self-tolerance. abstract of manuscript: Histone Deacetylase 7 (HDAC7) is a TCR signal-dependent regulator of differentiation that is highly expressed in CD4/CD8 double-positive (DP) thymocytes. Here we examine the effect of blocking TCR-dependent nuclear export of HDAC7 during thymic selection, through expression of a signal-resistant mutant of HDAC7 (HDAC7-?P) in thymocytes. We find that HDAC7-?P Transgenic thymocytes exhibit a profound block in negative thymic selection, but can still undergo positive selection, resulting in the escape of autoreactive T cells into the periphery. Gene expression profiling reveals a comprehensive suppression of the negative selection-associated gene expression program in DP thymocytes, associated with a defect in the activation of MAP kinase pathways by TCR signals. The consequence of this block in vivo is a lethal autoimmune syndrome involving the exocrine pancreas and other abdominal organs. These experiments establish a novel molecular model of autoimmunity and cast new light on the relationship between thymic selection and immune self-tolerance. Goal of Microarray experiment: We did these experiments to determine how alteration of the function of HDAC7, a site-specific and signal-dependent repressor of transcription, changes gene expression in CD4/CD8 DP thymocytes.
Histone deacetylase 7 regulates cell survival and TCR signaling in CD4/CD8 double-positive thymocytes.
Specimen part, Treatment
View SamplesThe transcriptome of P. aeruginosa PAO1 in the presence of extracelluar 2-oxoglutarate at a concentration of 20 mM.
Genetic analysis of the assimilation of C5-dicarboxylic acids in Pseudomonas aeruginosa PAO1.
No sample metadata fields
View SamplesAn early settlement of a complex gut microbiota can protect against gastro-intestinal dysbiosis, but the effects of neonatal microbiota colonization and early life feeding of medium chain triglycerides on the maturation of the porcine gastric mucosa are largely unknown.
The effects of starter microbiota and the early life feeding of medium chain triglycerides on the gastric transcriptome profile of 2- or 3-week-old cesarean delivered piglets.
Specimen part
View SamplesAn early settlement of a complex gut microbiota can protect against gastro-intestinal dysbiosis, but the effects of neonatal microbiota colonization on the maturation of the porcine gastric mucosa are largely unknown.
The effects of starter microbiota and the early life feeding of medium chain triglycerides on the gastric transcriptome profile of 2- or 3-week-old cesarean delivered piglets.
Specimen part
View SamplesThe intracellular pathogen Trypanosoma cruzi secretes an activity that blocks TGF--dependent induction of connective tissue growth factor (CTGF/CCN2). Here, we address the mechanistic basis for T. cruzi-mediated interference of
A soluble factor from Trypanosoma cruzi inhibits transforming growth factor-ß-induced MAP kinase activation and gene expression in dermal fibroblasts.
Specimen part
View SamplesSome aspects of the gene expression-based classification method were robust because the gliomasphere cultures retained their classification over many passages, and IDH1 mutant gliomaspheres were all proneural. While gene expression of a subset of gliomasphere cultures was more like the parent tumor than any other tumor, gliomaspheres did not always harbor the same classification as their parent tumor. Classification was not associated with whether a sphere culture was derived from primary or recurrent GBM or associated with the presence of EGFR amplification or rearrangement. Unsupervised clustering of gliomasphere gene expression distinguished 2 general categories (mesenchymal and nonmesenchymal), while multidimensional scaling distinguished 3 main groups and a fourth minor group. Unbiased approaches revealed that PI3Kinase, protein kinase A, mTOR, ERK, Integrin, and beta-catenin pathways were associated with in vitro measures of proliferation and sphere formation. Associating gene expression with gliomasphere phenotypes and patient outcome, we identified genes not previously associated with GBM: PTGR1, which suppresses proliferation, and EFEMP2 and LGALS8, which promote cell proliferation.
Large-scale assessment of the gliomasphere model system.
Disease
View SamplesBACKGROUND & AIMS- More frequent interaction of bacteria with the colonic epithelium is associated with ulcerative colitis (UC). The identities of all proteins which promote bacterial clearance in colonic epithelial cells are unknown. Previously, we discovered that dCAP-D3 (Chromosome Associated Protein-D3), regulates responses to bacterial infection. We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS- Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing CAP-D3 shRNA. CAP-D3 levels in colonic epithelial cells from healthy and UC patient tissues were analyzed by immunoblot. RNA-sequencing identified bacterially-induced CAP-D3 target genes. The role of CAP-D3 target genes in bacterial clearance was analyzed by gentamycin protection assays, immunofluorescent staining, and by using pharmacologic inhibitors. RESULTS- CAP-D3 expression was reduced in colonic epithelial cells from UC patients with active disease. Reduction of CAP-D3 expression inhibited autophagy and decreased intracellular bacterial clearance. The components of the heterodimeric SLC7A5/SLC3A2 amino acid transporter were identified as CAP-D3 target genes; their levels increased in infected, CAP-D3 deficient cell lines and in cells from UC patients. In HT-29 cells, this resulted in earlier SLC7A5 recruitment to Salmonella-containing vacuoles, increased mTOR activity, and enhanced bacterial survival. Inhibition of SLC7A5/SLC3A2 or mTOR activity rescued the bacterial clearance defect in CAP-D3 deficient cells. CONCLUSIONS- CAP-D3 attenuates amino acid transporter transcription to promote bacterial autophagy in colon epithelial cells. CAP-D3 protein levels are decreased in patients with active UC, suggesting that CAP-D3 is a potential therapeutic target to restore mucosal homeostasis in UC patients. Overall design: Three RNA samples from 3 independent experiments including timepoints taken at 0, 0.5 and 7 hours post-infection were analyzed on a bioanalyzer for quality; one of the 0.5 hour post-infection samples was excluded at this time due to poor RNA purity. Directional, cDNA libraries made from cellular mRNAs were generated from the other 8 samples and sequenced (paired-end sequencing of 100 bp reads) in the Genomics Core at the University of Chicago on an Illumina HiSeq2000.
Chromosome-associated protein D3 promotes bacterial clearance in human intestinal epithelial cells by repressing expression of amino acid transporters.
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