Purpose: To identify genes that are transcriptionally controlled by Notch signaling during zebrafish lateral line proneuromast formation. Methods: We isolated primordium cells from dissected tails of 36 hpf Tg((cldnB:GFP);Tg(cldnB:gal4) x Tg(UAS:nicd)) and sibling Tg((cldnB:GFP);Tg(cldnB:gal4)) embryos by FACS and performed RNASeq analysis. Results: Using an optimized data analysis workflow, we mapped about 26 million sequence reads per sample to the zebrafish genome (build danRer10) and identified 32,105 transcripts in the dissociated tails of WT and NICD zebrafish with TopHat workflow. Approximately 2% of the transcripts showed differential expression between the WT and NICD tails, with a fold change =0.5 and p value <0.01. Conclusion: RNASeq analyses revealed that Notch signaling cell-autonomously induces apical constriction and cell adhesion. Overall design: Zebrafish lateral line mRNA profiles of 36 hours wild type (WT) and NICD embryos were generated in triplicate, using HiSeq 2500 (Illumina).
Proliferation-independent regulation of organ size by Fgf/Notch signaling.
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The quantitative deep bulk MARS-seq analysis demonstrated that DCs from ICAM-1/2 double knockout (DKO) chimeric LNs display similar transcriptomes to those of WT DCs in both their resting and CD40 mAb activated states. Overall design: Transciptome analysis of activated and resting classical DCs from either WT or ICAM-1/2 DKO chimeric mice was performed. DC cells were isolated from popliteal lymph nodes and sorted according to the following markers: CD45, CD11c and MHC-II
ICAMs Are Not Obligatory for Functional Immune Synapses between Naive CD4 T Cells and Lymph Node DCs.
Specimen part, Cell line, Treatment, SubjectView Samples
Swiss-Webster female mice (Charles River Laboratories, Wilmington, MA) 5-6 weeks of age were infected intranasally with 5 LD50 of either WT or lpp mutant of Y. pestis CO92. Uninfected mice were used as controls. At either 12 or 48 h post infection (p.i.), 3 mice per group were euthanized and the lungs, livers, and spleens were harvested and homogenized in 1 ml of RNALater (Ambion/Applied Biosystems, Austin, TX) using 50-ml tissue homogenizers (Kendell, Mansfield, MA). RNA was isolated from the tissue homogenates and purified using RNAqueous (Ambion). After an overnight precipitation, the RNA was resuspended in 20 ul of diethylpyrocarbonate (DEPC)-treated water and hybridized to Affymetrix GeneChip Mouse Genome 430 2.0 arrays, performed by the Molecular Genomics Core at UTMB Galveston, Texas, per manufacture protocols. The arrays had 45,000 probe sets representing more than 39,000 transcripts derived from ~34,000 well-substantiated mouse genes. The experiments were performed in triplicate (biological replicates), generating a total of 45 arrays.
Comparative Analyses of Transcriptional Profiles in Mouse Organs Using a Pneumonic Plague Model after Infection with Wild-Type Yersinia pestis CO92 and Its Braun Lipoprotein Mutant.
Sex, Specimen part, TimeView Samples
Analysis of H292 cells infected with Mycoplasma hyorhinis. Mycoplasma infection reduces the cytotoxic effect of Nutlin3 on H292 cells. The results provide insight into molecular mechanisms underlying the response of H292 cells to Nutlin3.
Mycoplasma hyorhinis reduces sensitivity of human lung carcinoma cells to Nutlin-3 and promotes their malignant phenotype.
Specimen part, Cell lineView Samples
SETD5 gene mutations have been identified as a frequent cause of idiopathic intellectual disability. Here we show that Setd5 haploinsufficient mice present developmental defects such as abnormal brain to body weight ratio and neural crest defect associated phenotypes. Furthermore, Setd5 mutant mice show impairments in cognitive tasks, enhanced long-term potentiation, delayed ontogenetic profile of ultrasonic vocalisation and behavioural inflexibility. Behavioural issues are accompanied by abnormal expression of postsynaptic density proteins previously associated with cognition. Our data suggest that Setd5 might regulate RNA polymerase II dynamics and gene transcription during development and learning via its interaction with the Hdac3 and Paf1 complexes. Our results emphasize the decisive role of Setd5 in a biological pathway found to be disrupted in intellectual disability and autism spectrum disorder patients. Overall design: RNA-sequencing for wild type and Setd5 heterozygous knockout mice in two settings. First, in whole embryo samples (age E9.5), three biological replicates each. Second, gene expression changes due to contextual fear conditioning (CFC) was studied by comparing baseline transcription in homecage (HC) mice with transcription one hour (CFC_1h) or three hours (CFC_3h) after fear conditioning (4-5 biological replicates per time point and genotype).
Haploinsufficiency of the intellectual disability gene SETD5 disturbs developmental gene expression and cognition.
Specimen part, Cell line, SubjectView Samples
To determine the regulatory pathways necessary for astrocytoma formation within complex adult brain microenvironments, we engineered mice for adult astrocyte-specific disruption of key regulators (pRb, Kras and Pten). Drivers of all astrocytoma grades were identified using CreERTM-inducible alleles. Inactivation of pRb was necessary to initiate grade II disease, and was the only lesion to do so. Additional activation of Kras progressed disease to grade III, while further Pten inactivation facilitated grade IV (glioblastoma) progression. These outcomes were elicited whether somatic events were induced broadly or focally. In vivo inactivation of pRb, which induced astrocyte proliferation and apoptosis, activated the MAPK pathway, while Kras activation and Pten loss triggered PI3K pathways.
Evolutionary etiology of high-grade astrocytomas.
Sex, TimeView Samples
Analysis of global gene expression in myeloid cells infiltrating tumors after irradiation. Cell death induces recruitment of myeloid cells into irradiated tumors thereby stimulating tumor recurrence. Results provide insights into molecular mechanisms regulating tumorigenic functions of myeloid cells in tumors re-growing after radiation therapy. Overall design: Samples were collected at day 4 from irradiated tumors in WT, TLR9KO and Stat3KO (MxCre/Stat3flox). There were total 11 samples with Â 3-4 replicates of each sample type.
TLR9 signaling in the tumor microenvironment initiates cancer recurrence after radiotherapy.
Specimen part, SubjectView Samples
Alveolar macrophages orchestrate pulmonary innate immunity and are essential for early immune surveillance and clearance of microorganisms in the airways. Inflammatory signaling must be sufficiently robust to promote host defense but limited enough to prevent excessive tissue injury. Macrophages in the lungs utilize multiple transcriptional and post-transcriptional mechanisms of inflammatory gene expression to delicately balance the elaboration of immune mediators. RNA terminal uridyltransferases (TUTs), including the closely homologous family members Zcchc6 (TUT7) and Zcchc11 (TUT4), have been implicated in the post-transcriptional regulation of inflammation from studies conducted in vitro. In vivo, we observed that Zcchc6 is expressed in mouse and human primary macrophages. Zcchc6-deficient mice are viable and born in Mendelian ratios and do not exhibit an observable spontaneous phenotype under basal conditions. Following an intratracheal challenge with S. pneumoniae, Zcchc6 deficiency led to a modest but significant increase in the expression of select cytokines including IL-6, CXCL1, and CXCL5. These findings were recapitulated in vitro whereby Zcchc6-deficient macrophages exhibited similar increases in cytokine expression due to bacterial stimulation. Although loss of Zcchc6 also led to increased neutrophil emigration to the airways during pneumonia, these responses were not sufficient to impact host defense against infection.
The RNA uridyltransferase Zcchc6 is expressed in macrophages and impacts innate immune responses.
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To evaluate the transcriptomes of lesional skin from different body parts of the same individual. Specifically, we conducted a transcriptomic study to investigate expression variability for diseased samples taken from different anatomic regions of same patient, and to compare the variability to between individuals variability. Overall design: 5 psoriasis patients, each with 4 psoriatic and 1 uninvolved skin biopsies. Totally 25 RNA-seq experiments conducted.
Transcriptional determinants of individualized inflammatory responses at anatomically separate sites.
Specimen part, Disease stage, SubjectView Samples