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SRP064266
Mus musculus
12 Downloadable Samples
Illumina HiSeq 2000
Description
YY1 is a ubiquitously expressed transcription factor that has been demonstrated to be essential for pro-B cell development. However, the role of YY1 in other B cell populations has never been investigated. It has been proposed that YY1 is a key regulator for the germinal center B cell program since the YY1 motif was present in much higher frequency in germinal center B cell signature genes than signature genes of other B cell subsets. Indeed, in accord with this prediction, we demonstrated that deletion of YY1 by Cg1-Cre completely prevented differentiation of naïve B cells into germinal center B cells and plasma cells after antigen stimulation. To determine if YY1 was also required for the differentiation of other B cell populations, we deleted YY1 with CD19-Cre and found that all peripheral B cell subsets including B1 B cells require YY1 for their differentiation. By deleting YY1 acutely with ER-Cre, we demonstrated that all B cell subsets require YY1 for their maintenance. ChIP-seq shows that YY1 predominantly binds to promoters, and pathway analysis of the genes which bind YY1 show that they are enriched in ribosomal functions, mitochondrial functions such as bioenergetics, and functions related to transcription, such as mRNA splicing, metabolism of RNA. By RNA-seq analysis of differentially expressed genes, we demonstrated that YY1 normally activates genes involved in mitochondrial bioenergetics, while it normally downregulates genes involved in transcription, mRNA splicing, NF-kB signaling pathways, AP-1 transcription factor network, chromatin remodeling, cytokine signaling pathways, cell adhesion, cell proliferation and c-Myc targets. Overall design: Total RNA was prepared from RAG-/-pro-B cells, RAG-/-YY1f/f x mb1-Cre pro-B cells, RAG-/- µ+ pre-B cells, C57BL/6 follicular B cells, and C57BL/6 GC B cells. RNA was extracted using TRIzol (Life Technologies) and genomic DNA was eliminated using the genomic DNA wipeout buffer in the QuantiTect Reverse transcription kit (Qiagen). A final purification of the RNA was performed with the RNeasy kit (Qiagen). Ribosomal RNA was eliminated using Ribo-Zero Magnetic Gold Kit (Illumina).RNA samples were submitted to the Next Generation Sequencing Core, where they were processed with the NEBNext Ultra Directional RNA Library Prep Kit for Illumina and sequenced on the Illumina HiSeq. Three independent RNA-seq samples were used for RAG-/- pro-B and RAG-/- YY1f/f x mb1-Cre pro-B cells, and two samples for the other cell types.
Publication Title
YY1 plays an essential role at all stages of B-cell differentiation.
Sample Metadata Fields
Sex, Specimen part, Cell line, Subject
View Samples- Mus musculus
- 2 Downloadable Samples
- Illumina HiSeq 2000
Description
A diverse antibody repertoire is formed through the rearrangement of V, D, and J segments at the immunoglobulin heavy chain (Igh) loci. The C57BL/6 murine Igh locus has over 100 functional VH gene segments that can recombine to a rearranged DJH. While the non-random usage of VH genes is well documented, it is not clear what elements determine recombination frequency. To answer this question we conducted deep sequencing of 5’-RACE products of the Igh repertoire in pro-B cells, amplified in an unbiased manner. ChIP-seq results for several histone modifications and RNA polymerase II binding, RNA-seq for sense and antisense non-coding germline transcripts, and proximity to CTCF and Rad21 sites were compared to the usage of individual V genes. Computational analyses assessed the relative importance of these various accessibility elements. These elements divide the Igh locus into four epigenetically and transcriptionally distinct domains, and our computational analyses reveal different regulatory mechanisms for each region. Proximal V genes are relatively devoid of active histone marks and non-coding RNA in general, but having a CTCF site near their RSS is critical, suggesting that position near the base of the chromatin loops is important for rearrangement. In contrast, distal V genes have high levels of histone marks and non-coding RNA, which may compensate for their poorer RSS and for being distant from CTCF sites. Thus, the Igh locus has evolved a complex system for the regulation of V(D)J rearrangement that is different for of each the four domains that comprise this locus. Overall design: RNA was extracted from C57BL/6 RAG-/- pro-B cells using Trizol® (Life Technologies Corp., Carlsbad CA) and genomic DNA was eliminated using the genomic DNA wipeout buffer in the QuantiTect Reverse transcription kit (QIAGEN). A final purification of the RNA was performed with the RNeasy kit from QIAGEN. For each sample, 100 ng of total RNA was used to make RNASeq libraries using the NuGEN Encore Complete DR kits following manufacturer''s recommended protocols. Sequencing libraries were gel purified to ensure insert sizes were larger than 100 bp in length and sequenced on an Ilumina HiSeq2000 for 100 bases plus 7 bases for indexing.
Publication Title
Deep sequencing of the murine IgH repertoire reveals complex regulation of nonrandom V gene rearrangement frequencies.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Homo sapiens
- 38 Downloadable Samples
- Illumina HiSeq 3000
Description
Understanding how differentiation, microenvironment, and hormonal milieu influence human breast cell susceptibility to malignant transformation will require the use of physiologically relevant in vitro systems. We developed a 3D culture model that enables the propagation of normal estrogen receptor alpha (ER)+ cells. The purpose of this experiment was to assess ER functionality and compare estrogen-induced transcripts among samples and systems. Overall design: RNA-seq was performed on RNA prepared from replicate 3D cultures from 3 normal 3D breast culture specimens exposed to 10nM estradiol or vehicle alone for 6 or 24 hours.
Publication Title
Propagation of functional estrogen receptor positive normal human breast cells in 3D cultures.
Sample Metadata Fields
Specimen part, Treatment, Subject, Time
View Samples- Mus musculus
- 94 Downloadable Samples
Affymetrix Murine Genome U74A Version 2 Array (mgu74av2)
Description
Pneumocystis is a pathogen of immunocompromised hosts but can also infect healthy hosts, in whom infection is rapidly controlled and cleared. To better understand the immune mechanisms contributing to clearance of infection, microarray methods were used to examine differential gene expression in the lungs of C57BL/6 and CD40 ligand knock-out (CD40L-KO) mice over time following exposure to Pneumocystis. Immuncompetent C57BL/6 mice, which control and clear infection efficiently, showed a robust response to infection characterized by the upregulation of 349 primarily immune-response associated genes. Temporal changes in the expression of these genes suggested that there was an early (week 2) primarily innate response, that waned without controlling infection; this were followed by primarily adaptive immune responses that peaked at week 5 and successfully cleared the infection. In conjunction with the latter, there was an increased expression of B cell associated (immunoglobulin) genes at week 6 that persisted through 11 weeks. In contrast, CD40L-KO mice, which are highly susceptible to developing severe Pneumocystis pneumonia, showed essentially no upregulation of immune-response associated genes at days 35 to 75. Immunohistochemical staining supported these observations by demonstrating an increase in CD4+, CD68+, and CD19+ cells in C57BL/6 but not CD40L-KO mice. Thus, the healthy host demonstrates a robust biphasic response to infection by Pneumocystis; CD40 ligand is an essential upstream regulator of the adaptive immune responses that efficiently control infection and prevent development of progressive pneumonia.
Publication Title
Immune responses to Pneumocystis murina are robust in healthy mice but largely absent in CD40 ligand-deficient mice.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 8 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Background: Cysteinyl leukotrienes (cysLTs) are important mediators of innate immune responsiveness and chronic inflammatory diseases. CysLTs acting through cysteinyl leukotriene receptors may influence the migration and activity of cells such as eosinophils, monocytes and dendritic cells.
Publication Title
Leukotriene D(4) induces gene expression in human monocytes through cysteinyl leukotriene type I receptor.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 8 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
LTB4, 50 nmol/L for 30 minutes, induced expression of 27 genes in cultured human elutriated monocytes comparred to vehicle (ethanol) treated control cells.
Publication Title
Cooperative and redundant signaling of leukotriene B4 and leukotriene D4 in human monocytes.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Homo sapiens
- 26 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
In sickle cell disease, ischemia-reperfusion injury and intravascular hemolysis produce endothelial dysfunction and vasculopathy characterized by reduced nitric oxide (NO) and arginine bioavailability. Recent functional studies of platelets in patients with sickle cell disease reveal a basally activated state, suggesting that pathological platelet activation may contribute to sickle cell disease vasculopathy. Studies were therefore undertaken to examine transcriptional signaling pathways in platelets that may be dysregulated in sickle cell disease. We demonstrate and validate here the feasibility of comparative platelet transcriptome studies on clinical samples from single donors, by the application of RNA amplification followed by microarray-based analysis of 54,000 probe sets. Data mining an existing microarray database, we identified 220 highly abundant genes in platelets and a subset of 72 relatively platelet-specific genes, defined by more than 10-fold increased expression compared to the median of other cell types in the database with amplified transcripts. The highly abundant platelet transcripts found in the current study included 82% or 70% of platelet abundant genes identified in two previous gene expression studies on non-amplified mRNA from pooled or apheresis samples, respectively. On comparing the platelet gene expression profiles in 18 patients with sickle cell disease in steady state to 12 African American controls, at a 3-fold cut-off and 5% false discovery rate, we identified ~100 differentially expressed genes, including multiple genes involved in arginine metabolism and redox homeostasis. Further characterization of these pathways using real time PCR and biochemical assays revealed increased arginase II expression and activity and decreased platelet polyamine levels. These studies suggest a potential pathogenic role for platelet arginase and altered arginine and polyamine metabolism in sickle cell disease and provide a novel framework for the study of disease-specific platelet biology.
Publication Title
Amplified expression profiling of platelet transcriptome reveals changes in arginine metabolic pathways in patients with sickle cell disease.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 9 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
Changes in microRNA and mRNA expression with differentiation of human bronchial epithelial cells.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 9 Downloadable Samples
Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Normal human bronchial epithelial (NHBE) cells cultured in an air-liquid interface (ALI) system form a polarized, pseudostratified epithelium composed of basal, ciliated and goblet cells that closely resemble the in vivo airway epithelium structure. ALI cultures of NHBE cells provide a unique in vitro system to investigate airway epithelial biology, including developmental, structural and physiologic aspects. In this study, we wanted to investigate mRNA expression patterns during airway epithelium differentiation.
Publication Title
Changes in microRNA and mRNA expression with differentiation of human bronchial epithelial cells.
Sample Metadata Fields
Specimen part
View Samples- Rattus norvegicus
- 38 Downloadable Samples
- Affymetrix Rat Expression 230A Array (rae230a)
Description
The expression levels of many genes show wide natural variation among strains or populations. This study investigated the potential for animal strain-related genotypic differences to confound gene expression profiles in acute cellular rejection (ACR). Additional analysis allowed for selection of 49 candidate genes uniquely associated with ACR, but only after accounting for the unexpectedly large effect of animal strain. Studies of ACR that examine gene expression in peripheral blood may be confounded by strain differences. These results indicate the need for study designs that eliminate or control for the large effect of genetic background on the transcriptome of immune cells.
Publication Title
Impact of animal strain on gene expression in a rat model of acute cardiac rejection.
Sample Metadata Fields
Specimen part
View Samples