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SRP060247
Mus musculus
6 Downloadable Samples
Illumina HiSeq 2500
Description
Gamma-herpesviruses encode a cytoplasmic mRNA-targeting endonuclease, termed SOX, that cleaves the majority of mRNAs within a cell. Cleaved fragments are subsequently degraded by the cellular mRNA degradation machinery. Here, we reveal that mammalian cells respond to this widespread cytoplasmic mRNA decay by altering levels of RNA polymerase II (RNAPII) transcription in the nucleus. Measurements of both RNAPII recruitment to promoters and nascent mRNA synthesis revealed that the majority of affected genes are transcriptionally repressed in SOX-expressing cells. The transcriptional feedback does not occur in response to the initial endonuclease-induced cleavage, but instead to degradation of the cleaved fragments by cellular exonucleases. In particular, Xrn1 catalytic activity is required for transcriptional repression. Notably, viral mRNA transcription escapes decay-induced repression, and this escape requires Xrn1. Collectively, these results indicate that mRNA decay rates impact transcription in mammalian cells, and that gamma-herpesviruses have incorporated this feedback mechanism into their own gene expression strategy. Overall design: NIH 3T3 cells were mock, WT, or ?HS infected with MHV68 in duplicate and 4sU-labeled RNA isolated. 4sU-labeled RNA was submitted for sequencing and reads aligned to the mouse genome or MHV68 viral genome. Differential cellular gene expression was determined between mock and WT infected, mock and ?HS infected, as well as differential viral gene expression between WT and ?HS.
Publication Title
Viral Nucleases Induce an mRNA Degradation-Transcription Feedback Loop in Mammalian Cells.
Sample Metadata Fields
No sample metadata fields
View Samples- Rattus norvegicus
- 6 Downloadable Samples
Affymetrix Rat Genome 230 2.0 Array (rat2302)
Description
SHH signaling pathway is activated in many type of cancers. However, the role of its activation in particular type of cancer was poorly understood. The GLI family transcription factor GLI1 is the effector of Shh pathway activation and functions as oncogene. Our goal of research is to identify the GLI1 targets in desmoplastic medulloblastomas.
Publication Title
Defining a role for Sonic hedgehog pathway activation in desmoplastic medulloblastoma by identifying GLI1 target genes.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 20 Downloadable Samples
- Affymetrix Human Genome U95 Version 2 Array (hgu95av2)
Description
Considerable variation in gene expression data from different DNA microarray platforms has been demonstrated. However, no characterization of the source of variation arising from labeling protocols has been performed. To analyze the variation associated with T7-based RNA amplification/labeling methods, aliquots of the Stratagene Human Universal Reference RNA were labeled using 3 eukaryotic target preparation methods and hybridized to a single array type (Affymetrix U95Av2). Variability was measured in yield and size distribution of labeled products, as well as in the gene expression results. All methods showed a shift in cRNA size distribution, when compared to un-amplified mRNA, with a significant increase in short transcripts for methods with long IVT reactions. Intra-method reproducibility showed correlation coefficients >0.99, while inter-method comparisons showed coefficients ranging from 0.94 to 0.98 and a nearly two-fold increase in coefficient of variation. Fold amplification for each method was positively correlated with the number of present genes. Two factors that introduced significant bias in gene expression data were observed: a) number of labeled nucleotides that introduces sequence dependent bias, and b) the length of the IVT reaction that introduces a transcript size dependent bias. This study provides evidence of amplification method dependent biases in gene expression data.
Publication Title
In vitro transcription amplification and labeling methods contribute to the variability of gene expression profiling with DNA microarrays.
Sample Metadata Fields
No sample metadata fields
View Samples- Saccharomyces cerevisiae
- 24 Downloadable Samples
- Illumina Genome Analyzer IIx
Description
Transfer RNA (tRNA) modifications enhance the efficiency, specificity and fidelity of translation in all organisms. The anticodon modification mcm5s2U34 is required for normal growth and stress resistance in yeast; mutants lacking this modification have numerous phenotypes. Mutations in the homologous human genes are linked to neurological disease. The yeast phenotypes can be ameliorated by overexpression of specific tRNAs, suggesting that the modifications are necessary for efficient translation of specific codons. We determined the in vivo ribosome distributions at single codon resolution in yeast strains lacking mcm5s2U. We found accumulations at AAA, CAA, and GAA codons, suggesting that translation is slow when these codons are in the ribosomal A site, but these changes appeared too small to affect protein output. Instead, we observed activation of the GCN4-mediated stress response by a non- canonical pathway. Thus, loss of mcm5s2U causes global effects on gene expression due to perturbation of cellular signaling. Overall design: WT yeast and mutants lacking anticodon tRNA modifications were grown in YPD, and subjected to ribosome footprint profiling (ribo-seq) and RNA-seq of poly-A selected RNA. Dataset contains biological replicates for WT, ?ncs6 and ?uba4. Technical replicates were also performed for all RNA-seq datasets (using a different poly-A selection method).
Publication Title
Loss of a conserved tRNA anticodon modification perturbs cellular signaling.
Sample Metadata Fields
Cell line, Subject
View Samples- Homo sapiens
- 16 Downloadable Samples
- Illumina HiSeq 2000
Description
Idiopathic pulmonary fibrosis (IPF) is a progressive disease of the middle aged and elderly with a prevalence of one million persons worldwide. The fibrosis spreads from affected alveoli into contiguous alveoli, creating a reticular network that leads to death by asphyxiation. Lung fibroblasts from patients with IPF have phenotypic hallmarks, distinguishing them from their normal counterparts: pathologically activated Akt signaling axis, increased collagen and a-smooth muscle actin expression, distinct gene expression profile, and ability to form fibrotic lesions in model organisms. Despite the centrality of these fibroblasts in disease pathogenesis, their origin remains uncertain. Here, we report the identification of cells in the lungs of patients with IPF with the properties of mesenchymal progenitors. In contrast to progenitors isolated from nonfibrotic lungs, IPF mesenchymal progenitor cells produce daughter cells manifesting the full spectrum of IPF hallmarks, including the ability to form fibrotic lesions in zebrafish embryos and mouse lungs, and a transcriptional profile reflecting these properties. Morphological analysis of IPF lung tissue revealed that mesenchymal progenitor cells and cells with the characteristics of their progeny comprised the fibrotic reticulum. These data establish that the lungs of patients with IPF contain pathological mesenchymal progenitor cells that are cells of origin for fibrosis-mediating fibroblasts. These fibrogenic mesenchymal progenitors and their progeny represent an unexplored target for novel therapies to interdict fibrosis. Overall design: RNA-seq of lung fibroblasts from IPF or healthy control patients at day 0 or day 21 of culture.
Publication Title
Calcium-binding protein S100A4 confers mesenchymal progenitor cell fibrogenicity in idiopathic pulmonary fibrosis.
Sample Metadata Fields
Specimen part, Disease, Disease stage, Subject
View Samples- Mus musculus
- 49 Downloadable Samples
- Illumina HiSeq 2000
Description
We previoiusly identified WDR11 as a potential tumor suppressor in murine medulloblastoma models. To determine additional genes/pathways affected by WDR11 overexpression.To compare somatic mutations of murine models with human medulloblastoma (MB), we performed whole-exome sequencing of mouse tumors representing three distinct MB subgroups: Wnt, Sonic Hedgehog (Shh) and Group 3 (G3). 64 somatic mutations were identified and validated, including 40 predicted to cause amino acid changes. After filtering and cross-species analysis with 366 human MBs from four independent studies, human orthologs for 16 of the 40 mouse genes were found to harbor non-silent mutations in human MB. Loss-of-function Mll2 mutations detected in one mouse tumor were previously reported in 30 of 366 human MBs. In mice with G3 MB, one mouse that died at least 15 days earlier than the others had four novel candidate genes harboring non-silent somatic mutations, Lrfn2, Smyd1, Ubn2 and Wdr11. To test whether these genes had tumor suppressive activity, we constitutively overexpressed each wild type gene in murine G3 tumorspheres followed by intracranial implantation. Mice harboring mouse G3 MB overexpressing WDR11 showed extended survival compared to the other three genes. Genes in the KEGG WNT signaling pathway, including Ccnd1/2/3, Myc and Tcf7l1, were down-regulated in G3 MB tumorspheres overexpressing WDR11, consistent with reduced tumor progression. In conclusion, we demonstrated that common spontaneous mutations were shared between human and murine models of MB suggesting similar molecular mechanisms of tumorigenesis, and identified WDR11 as a protein with tumor suppressive activity in G3 MB. Overall design: Compare differentially expressed genes in WDR11 overexpression group versus control group.
Publication Title
Exome sequencing analysis of murine medulloblastoma models identifies WDR11 as a potential tumor suppressor in Group 3 tumors.
Sample Metadata Fields
Specimen part, Treatment, Subject
View Samples- Populus trichocarpa
- 9 Downloadable Samples
- Affymetrix Poplar Genome Array (poplar)
Description
Embolism and the refilling of xylem vessels are intrinsic to the ability of plants to handle the transport of water under tension. While the formation of an embolized vessel is an abiotic process, refilling against the pressure gradient requires biological activity to provide both the energy and the water needed to restore xylem transport capacity.
Publication Title
Transcriptome response to embolism formation in stems of Populus trichocarpa provides insight into signaling and the biology of refilling.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Homo sapiens
- 8 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Genome instability is a potential limitation to the research and therapeutic application of induced pluripotent stem cells (iPSCs). Observed genomic variations reflect the combined activities of DNA damage, cellular DNA damage response (DDR), and selection pressure in culture. To understand the contribution of DDR on the distribution of copy number variations (CNVs) in iPSCs, we mapped CNVs of iPSCs with mutations in the central DDR gene ATM onto genome organization landscapes defined by genome-wide replication timing profiles. We show that following reprogramming the early and late replicating genome is differentially affected by CNVs in ATM deficient iPSCs relative to wild type iPSCs. Specifically, the early replicating regions had increased CNV losses during retroviral reprogramming. This differential CNV distribution was not present after later passage or after episomal reprogramming. Comparison of different reprogramming methods in the setting of defective DNA damage response reveals unique vulnerability of early replicating open chromatin to retroviral vectors.
Publication Title
Influence of ATM-Mediated DNA Damage Response on Genomic Variation in Human Induced Pluripotent Stem Cells.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 2 Downloadable Samples
- Illumina HiSeq 2000
Description
Transcriptome profiling using RNA-seq of ß-TC3 cell, a mouse pancreatic cell line used in the study of novel Cis-regulatory elements for the Pax6 gene . Overall design: Total RNA was collected and a Illumina sequencing libraries prepared from two biological replicates of cultured ß-TC3 cells.
Publication Title
Functional characteristics of novel pancreatic Pax6 regulatory elements.
Sample Metadata Fields
Cell line, Subject
View Samples- Mus musculus
- 10 Downloadable Samples
- Affymetrix Mouse Expression 430A Array (moe430a)
Description
Mice lacking p53 and one or two alleles of the cyclin D-dependent kinase inhibitor p18Ink4c are prone to medulloblastoma development. The tumor frequency is increased by exposing postnatal animals to ionizing radiation at a time when their cerebella are developing. In irradiated mice engineered to express a floxed p53 allele and a Nestin-Cre transgene, tumor development can be restricted to the brain. Analysis of these animals indicated that inactivation of one or both Ink4c alleles did not affect the time of medulloblastoma onset but increased tumor invasiveness. All such tumors exhibited complete loss of function of the Patched 1 (Ptc1) gene encoding the receptor for sonic hedgehog, and many exhibited other recurrent genetic alterations, including trisomy of chromosome 6, amplification of N-Myc, modest increases in copy number of the Ccnd1 gene encoding cyclin D1, and other complex chromosomal rearrangements. In contrast, medulloblastomas arising in Ptc1+/- mice lacking one or both Ink4c alleles retained p53 function and exhibited only limited genomic instability. Nonetheless, complete inactivation of the wild type Ptc1 allele was a universal event, and trisomy of chromosome 6 was again frequent. The enforced expression of N-Myc or cyclin D1 in primary cerebellar granule neuron precursors isolated from Ink4c-/-, p53-/- mice enabled the cells to initiate medulloblastomas when injected back into the brains of immunocompromised recipient animals. These engineered tumors exhibited gene expression profiles indistinguishable from those of medulloblastomas that arose spontaneously. These results underscore the functional interplay between a network of specific genes that recurrently contribute to medulloblastoma formation.
Publication Title
Genetic alterations in mouse medulloblastomas and generation of tumors de novo from primary cerebellar granule neuron precursors.
Sample Metadata Fields
No sample metadata fields
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