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SRP013912
Homo sapiens
4 Downloadable Samples
Illumina Genome Analyzer II
Description
MicroRNAs (miRNAs) are a class of small RNA molecules that regulate expression of specific mRNA targets. They can be released from cells, often encapsulated within extracellular vesicles (EVs), and therefore have the potential to mediate intercellular communication. It has been suggested that certain miRNAs may be selectively exported, although the mechanism has yet to be identified. Manipulation of the miRNA content of EVs will be important for future therapeutic applications. We therefore wished to assess which endogenous miRNAs are enriched in EVs and how effectively an overexpressed miRNA would be exported. Small RNA libraries from HEK293T cells and vesicles before or after transfection with a vector for miR-146 overexpression were analysed by deep sequencing. A subset of miRNAs was found to be enriched in EVs. The global expression data provided by deep sequencing confirms that specific miRNAs are enriched in EVs released by HEK293T cells. Overall design: Cells were transfected with a plasmid to direct overexpression of miR-146a. Extracellular vesicles were isolated by ultracentrifugation from untreated and transfected cells. RNA was isolated from one sample each of untreated and transfected cells and vesicles.Small RNA libraries were prepared for sequencing.
Publication Title
Selective extracellular vesicle-mediated export of an overlapping set of microRNAs from multiple cell types.
Sample Metadata Fields
Specimen part, Subject
View Samples- Rattus norvegicus
- 9 Downloadable Samples
- IlluminaGenomeAnalyzer
Description
MicroRNAs (miRNAs) are single-stranded non-coding RNAs that negatively regulate target gene expression through mRNA cleavage or translational repression. There is mounting evidence that they play critical roles in heart disease. The expression of known miRNAs in the heart has been studied at length by microarray and quantitative PCR but it is becoming evident that microRNA isoforms (isomiRs) are potentially physiologically important. It is well known that left ventricular (patho)physiology is influenced by transmural heterogeneity of cardiomyocyte phenotype, and this likely reflects underlying heterogeneity of gene expression. Given the significant role of miRNAs in regulating gene expression, knowledge of how the miRNA profile varies across the ventricular wall will be crucial to better understand the mechanisms governing transmural physiological heterogeneity. To determinine miRNA/isomiR expression profiles in the rat heart we investigated tissue from different locations across the left ventricular wall using deep sequencing. We detected significant quantities of 145 known rat miRNAs and 68 potential novel orthologs of known miRNAs, in mature, mature* and isomiR formation. Many isomiRs were detected at a higher frequency than their canonical sequence in miRBase and have different predicted targets. The most common miR-133a isomiR was more effective at targeting a construct containing a sequence from the gelsolin gene than was canonical miR-133a, as determined by dual-fluorescence assay. We identified a novel rat miR-1 homolog from a second miR-1 gene; and a novel rat miRNA similar to miR-676. We also cloned and sequenced the rat miR-486 gene which is not in miRBase (v18). Signalling pathways predicted to be targeted by the most highly detected miRNAs include Ubiquitin-mediated Proteolysis, Mitogen-Activated Protein Kinase, Regulation of Actin Cytoskeleton, Wnt signalling, Calcium Signalling, Gap junctions and Arrhythmogenic Right Ventricular Cardiomyopathy. Most miRNAs are not expressed in a gradient across the ventricular wall, with exceptions including miR-10b, miR-21, miR-99b and miR-486. Overall design: The hearts of 3 male 8 month old Sprague-Dawley rats were rapidly extracted after euthanasia with sodium pentobarbital. A section of the free wall of the left ventricle was dissected into epicardium, mid-myocardium and endocardium by cutting approximately 1 mm from the epicardial and endocardial surfaces. Small RNA was extracted (miRNeasy Kit; Qiagen, Crawley UK), quantified (Nanodrop; Thermo Scientific) and quality assessed for degradation (RNA Nano Chip, Bioanalyser 2100; Aligent Technologies, Wokingham UK; only samples with a RNA integrity no. (RIN) =8 were carried forward) and retention of small RNA (Small RNA Chip, Bioanalyser 2100). Small RNA was preferentially ligated with adapters, reverse transcribed into cDNA and amplified with 9 individually tagged primer indices (TruSeq Small RNA Sample Preparation Kit; Illumina, Little Chesterford, UK) and a library of small RNA created for each sample. After gel purification the cDNA products were again analysed on the bioanalyser using a High Sensitivity DNA Chip and assessed for the presence and concentration of the peak corresponding to ligated and tagged miRNA (approximately 147nt). Only samples with suitable RIN values exhibiting good retention of small RNA species were used for library preparation. After pooling, the samples were sequenced by TrinSeq (Trinity Genome Sequencing Lab & Neuropsychiatric Genetics Group, Trinity College Dublin, Ireland (http://www.medicine.tcd.ie/sequencing); using TruSeq SR Cluster Kit v5 (Illumina) and the resultant data trimmed and aligned to miRBase v18 (CLC Genomics Workbench v4.0; CLC bio, Swansea UK).
Publication Title
Distinctive profile of IsomiR expression and novel microRNAs in rat heart left ventricle.
Sample Metadata Fields
No sample metadata fields
View Samples- Bos taurus
- 1 Downloadable Sample
- Illumina Genome Analyzer II
Description
The aim of this study was to identify and quantify microRNAs and other small regulatory RNAs expressed in primary retinal microvascular endothelial cells (RMECs) using deep sequencing. RMECs were isolated, RNA extracted, a small RNA library prepared and deep sequencing performed. A total of 6.8 million reads were mapped to 250 known microRNAs in miRBase (release 16). Several novel microRNAs and multiple new members of the miR-2284/2285 family were detected. Several ~30 nucleotide sno-miRNAs were identified, with the most highly expressed being derived from snoRNA U78. Highly expressed microRNAs previously associated with endothelial cells included miR-126 and miR-378, but the most highly expressed was miR-21, comprising more than one third of all mapped reads. The independence from prior sequence knowledge provided by deep sequencing facilitates analysis of novel microRNAs and other small RNAs. This approach also enables quantitative evaluation of microRNA expression, which has highlighted the predominance of a small number of microRNAs in RMECs. Further characterisation of the functions of the highly expressed microRNAs will provide insights into endothelial biology. Overall design: Single sample of primary cell culture
Publication Title
Deep sequencing reveals predominant expression of miR-21 amongst the small non-coding RNAs in retinal microvascular endothelial cells.
Sample Metadata Fields
Specimen part, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Fetal and adult -globin gene expression is tightly regulated during human development. Fetal globin genes are transcriptionally silenced during embryogenesis through the process of hemoglobin switching. Efforts to understand the transcriptional mechanism(s) behind fetal globin silencing have led to novel strategies to derepress fetal globin expression in the adult, which could alleviate symptoms in hereditary b-globin disorders including sickle cell disease (SCD) and -thalassemia. We identified a novel zinc finger protein, pogo transposable element with zinc finger domain (Pogz), expressed in mouse and human hematopoietic stem and progenitor cells, which represses embryonic b-like globin gene expression in mice. Ablation of Pogz expression in adult hematopoietic cells in vivo results in persistence of embryonic b-like globin expression without significantly affecting erythroid development or mouse survival. Elevated embryonic -like globin expression correlates with reduced expression of Bcl11a, a known repressor of embryonic -like globin expression, in Pogz-/- fetal liver cells. Pogz binds to the Bcl11a promoter, and, to erythroid specific intragenic regulatory regions. Importantly, Pogz+/- mice develop normally, but show elevated embryonic b-like globin expression in peripheral blood cells, demonstrating that reducing Pogz levels results in persistence of embryonic b-like globin expression. Finally, knockdown of POGZ in primary human CD34+ hematopoietic stem and progenitor cell derived erythroblasts, reduces BCL11A expression and increases fetal hemoglobin expression. These findings are significant since new therapeutic targets and strategies are needed to treat the increasing global burden of b-globin disorders.
Publication Title
POGZ Is Required for Silencing Mouse Embryonic β-like Hemoglobin and Human Fetal Hemoglobin Expression.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 12 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
We compared the differentially expressed genes between the F9 Wt cells and F9 RAR gamma knock out cells before and after RA treatment. 3 replicates for each conditions.
Publication Title
Gene expression profiling elucidates a specific role for RARgamma in the retinoic acid-induced differentiation of F9 teratocarcinoma stem cells.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 31 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Primary glioblastoma, representing over 90% of adult glioblastoma, develop rapidly without preexisting lower-grade glioma. We have generated a mouse model of primary glioblastoma driven by a single p53 mutation. These p53-mutant gliomas lose the syntenic region of human chromosome 10q, which is mapped to mouse chr19 and chr7. Loss of mouse chr19, containing Pten, activates PI3K/Akt signaling.
Publication Title
Opposing Tumor-Promoting and -Suppressive Functions of Rictor/mTORC2 Signaling in Adult Glioma and Pediatric SHH Medulloblastoma.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 18 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
The goal of this study was to identify genes that are differentially expressed after genetic deletion of both alleles of the Cyp26a1 gene in murine embryonic stem cells. Cyp26a1 codes for the CYP26A1 enzyme which metabolizes RA to polar RA metabolites, such as 4-oxo-RA and 4-OH-RA. CYP26A1-/- ES cells do not metabolize RA within 48 hours of RA treatment while in Wt ES cells, polar RA metabolites are already detectable by 8 hr. In addition, the absence of CYP26A1 enzyme increases intracellular RA levels. By gene microarray analysis, we wanted to identify genes that would be affected by the lack of the Cyp26a1 gene.
Publication Title
CYP26A1 knockout embryonic stem cells exhibit reduced differentiation and growth arrest in response to retinoic acid.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 12 Downloadable Samples
- IlluminaHiSeq2000
Description
To gain insight into the molecular changes during OSCC carcinogenesis, we performed unbiased, whole genome deep sequencing (RNA-seq) using RNA isolated from cultured, human TERT-immortalized, non-tumorigenic OKF6-TERT1R and OSCC SCC-9 cells. Overall design: OKF6-TERT1R cells and SCC-9 cells were plated in 10 cm2 tissue culture plates at the density of 2 × 106 cells/plate and treated with 1 µM RA or vehicle (0.1% ethanol) for 48 hours. Experiment includes 3 independent biological replicates.
Publication Title
Altered histone mark deposition and DNA methylation at homeobox genes in human oral squamous cell carcinoma.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 12 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Retinoic acid receptors (RARs) , and are key regulators of embryonic development. Hematopoietic differentiation is regulated by RAR, and several types of leukemia show aberrant RAR activity. We demonstrate that RAR plays an important role in cellular memory and imprinting by regulating the CpG methylation status of specific promoter regions.
Publication Title
Epigenetic regulation by RARα maintains ligand-independent transcriptional activity.
Sample Metadata Fields
Cell line, Treatment
View Samples- Mus musculus
- 44 Downloadable Samples
- Illumina HiSeq 2000
Description
The complete transcriptomes of kidney cortex from 3 ?-HIF2aM3 18 month old TG+ male mice and 3 age matched wild type (WT) C57BL/6 male mice were sequenced on an Illumina HiSeq2000 Sequencer. Overall design: Examination of complete transcriptome of kidney cortex between ?-HIF2aM3 TG+ male mice and wild type C57BL/6 male mice
Publication Title
Activation of HIF2α in kidney proximal tubule cells causes abnormal glycogen deposition but not tumorigenesis.
Sample Metadata Fields
Sex, Specimen part, Cell line, Subject
View Samples