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SRP052229
Homo sapiens
42 Downloadable Samples
IlluminaHiSeq2000
Description
Roberts syndrome (RBS) is a human developmental disorder caused by mutations in the cohesin acetyltransferase ESCO2. We previously reported that mTORC1 was inhibited and overall translation was reduced in RBS cells. Treatment of RBS cells with L-leucine partially rescued mTOR function and protein synthesis, correlating with increased cell division. In this study, we use RBS as a model for mTOR inhibition and analyze transcription and translation with ribosome profiling to determine genome-wide effects of L-leucine. The translational efficiency of many genes is increased with Lleucine in RBS cells including genes involved in ribosome biogenesis, translation, and mitochondrial function. snoRNAs are strongly upregulated in RBS cells, but decreased with L-leucine. Imprinted genes, including H19 and GTL2, are differentially expressed in RBS cells consistent with contribution to mTORC1 control. This study reveals dramatic effects of L-leucine stimulation of mTORC1 and supports that ESCO2 function is required for normal gene expression and translation. Overall design: 42 samples of human fibroblast cell lines with various genotypes (wt, corrected, and esco2 mutants) are treated with l-leucine or d-leucine (control) for 3 or 24 hours. Biological replicates are present.
Publication Title
Improved transcription and translation with L-leucine stimulation of mTORC1 in Roberts syndrome.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 6 Downloadable Samples
- Illumina HiSeq 2500
Description
Cohesinopathies are characterized by mutations in the cohesin complex. Mutations in NIPBL, a cohesin loader, result in Cornelia de Lange syndrome (CdLS). CdLS is a congenital genetic disorder distinguished by craniofacial dysmorphism, abnormal upper limb development, delayed growth, severe cognitive retardation, and multiple organ malformations.It has been suggested that CdLS is caused by defects in the cohesin network that alter gene expression and genome organization. However, the precise molecular etiology of CdLS is largely unclear. To gain insights, we sequenced mRNAs isolated from mouse embryonic fibroblasts of both WT and NIPBL-haploinsufficient mice and compared their transcriptomes. Overall design: Examination of gene expression of WT and NIPBL+/- mice by RNA-seq
Publication Title
NIPBL Controls RNA Biogenesis to Prevent Activation of the Stress Kinase PKR.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 4 Downloadable Samples
- Illumina HiSeq 2500
Description
Condensin complexes are highly conserved for chromosome compaction to ensure their faithful segregation in mitosis. Condensin II is present in the nucleus throughout the cell cycle, including interphase. The aim of these experiments is to investigate the changes of gene expression in knockdown of NCAPH2, a condensin II subunit, in mouse embryonic stem cells compared to their control cells. Overall design: Examination of gene expression of controls and NCAPH2 knockdown cells by RNA-seq
Publication Title
Condensin II is anchored by TFIIIC and H3K4me3 in the mammalian genome and supports the expression of active dense gene clusters.
Sample Metadata Fields
Specimen part, Subject
View Samples- Danio rerio
- 45 Downloadable Samples
- IlluminaHiSeq2000
Description
Methods: Triplicate RNA samples from morphologically stage-matched embryos were sequenced to compare expression profiles over time. Strand-specific libraries were prepared using the TruSeq stranded total RNA-ribozero kit (Illumina) and 100-bp paired-end sequencing was performed to depth of 10 million reads per library on an Illumina HiSeq 2000. Methods: On average, 19 million 100 bp paired-end reads per library were generated. These were then adapter and quality trimmed using cutadapt and SolexaQA. Each sequencing data set was independently mapped to the zebrafish genome with a bowtie2 index generated from Danio_rerio.Zv9.70 (Ensembl) downloaded from Illumina's iGenomes collection. Zebrafish genome danRer7was used to provide known transcript annotations from Ensembl using TopHat2 (version 2.0.9) with the following options: “tophat2 --GTF genes.gtf --library-type fr-firststrand -p 24 --mate-inner-dist -8 --mate-std-dev 6 zv9” (on average, 75.38% reads mapped uniquely to the genome). Transcriptomes were assembled with Cufflinks (version 2.2.0) using options: 'cufflinks -p 32 --GTF genes.gtf' and differential expression analysis between control and knockdown embryos was performed using Cuffdiff. A FDR corrected p-value of 0.05 was applied as the cut off to identify differentially regulated transcripts Results: We could show that MO assisted depletion of Rad21 and CTCF affected the transcriptional profiles of embryos in different ways. Overall design: mRNA profiles of (2.5, 3.3, 4.5, 5.3, 10 hpf) wild type (WT) and morpholino depleted Rad21 MO (Rad21) and CTCF MO (CTCF) embryos were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000.
Publication Title
Cohesin facilitates zygotic genome activation in zebrafish.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 6 Downloadable Samples
Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
In euakryotes, mRNAs must be exported from the nucleus to the cytsoplasm. NXF2 is highly expressed in the mouse male germ cells. We are interested in its function in spermatogenesis, espically in the nuclear RNA export in the testis. To this end, we made Nxf2 mutant mice by gene targeting. In an attempt to identify the mRNA substrates of NXF2, we perform the microarray experiments on testes.
Publication Title
Inactivation of Nxf2 causes defects in male meiosis and age-dependent depletion of spermatogonia.
Sample Metadata Fields
No sample metadata fields
View Samples- Saccharomyces cerevisiae
- 27 Downloadable Samples
- Affymetrix Yeast Genome 2.0 Array (yeast2)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
The cohesin acetyltransferase Eco1 coordinates rDNA replication and transcription.
Sample Metadata Fields
No sample metadata fields
View Samples- Saccharomyces cerevisiae
- 27 Downloadable Samples
- Affymetrix Yeast Genome 2.0 Array (yeast2)
Description
Eco1 is an acetyltransferase subunit of the cohesin complex and acts during DNA replication to establish cohesion between sister chromatids. However, cohesin has additional functions in gene expression, DNA damage repair, and higher-order organization of chromosomes. The eco1 mutant W216G disrupts acetyltansferase activity, and causes genome-wide transcriptional defects which can be suppressed by deletion of FOB1, a gene also involved in DNA replication. This experiment investigates gene expression differences between the eco1-W216G mutant, and mutants in FOB1, and RAD61 a gene involved in inhibition of cohesion establishment but mutation of which is able to suppress temperature sensitivity of the eco1-W216G mutant.
Publication Title
The cohesin acetyltransferase Eco1 coordinates rDNA replication and transcription.
Sample Metadata Fields
No sample metadata fields
View Samples- Saccharomyces cerevisiae
- 35 Downloadable Samples
- Affymetrix Yeast Genome 2.0 Array (yeast2)
Description
The cohesin protein complex is well known for playing a role in chromosome segregation. However, it has additional less understood roles in transcription, DNA repair, and chromosome condensation. Mutants in two yeast orthologues (Eco1 and Scc2) of human cohesinopathy disease alleles were examined by transcriptional profiling in response to perturbation of the transcriptional program by amino acid starvation.
Publication Title
Cohesin proteins promote ribosomal RNA production and protein translation in yeast and human cells.
Sample Metadata Fields
Time
View Samples- Mus musculus
- 9 Downloadable Samples
- Illumina HiSeq 2500
Description
Cultured pluripotent stem cells are a cornerstone of regenerative medicine due to their ability to give rise to all cell types of the body. While pluripotent stem cells can be propagated indefinitely in vitro, pluripotency is paradoxically a very transient state in vivo, lasting 2-3 days around the time of blastocyst implantation. The exception to this rule is embryonic diapause, a reversible state of suspended development triggered by unfavorable conditions. Diapause is a strategy widely employed across the animal kingdom, including in mammals, but its regulation remains poorly understood. Here we report that inhibition of mechanistic target of rapamycin (mTor), a major nutrient sensor and promoter of growth, induces reversible pausing of mouse blastocyst development and allows their prolonged culture ex vivo. Paused blastocysts remain pluripotent and competent to give rise to embryonic stem (ES) cells and mice. We show that both natural diapause blastocysts in vivo and paused blastocysts ex vivo display pronounced reductions in mTor activity, translation and transcription. In addition, pausing can be induced directly in cultured ES cells and sustained for weeks in the absence of cell death or deviations from cell cycle distributions. We show that paused ES cells remain pluripotent, display a remarkable global suppression of transcription, and maintain a gene expression signature of diapaused blastocysts. These results allow for the first time the sustained suspension of development of a mammalian embryo in the laboratory, and shed light on the regulation of diapause and the origins of ES cells. Our findings have important implications in the fields of assisted reproduction, regenerative medicine, cancer, metabolic disorders and aging. Overall design: Examination of RNA expression profiles of embryonic stem cells in serum, 2i and paused states by RNA-seq
Publication Title
Inhibition of mTOR induces a paused pluripotent state.
Sample Metadata Fields
Specimen part, Cell line, Treatment, Subject
View Samples- Homo sapiens
- 9 Downloadable Samples
- Ion Torrent Proton
Description
Rationale - Regulatory T (Treg) cells suppress immune responses and have been shown to attenuate atherosclerosis. The Treg cell lineage specification factor FOXP3 is essential for Treg cells' ability to uphold immunological tolerance. In humans, FOXP3 exists in several different isoforms, however, their specific role is poorly understood. Objective - To define the regulation and functions of the two major FOXP3 isoforms, FOXP3fl and FOXP3?2, as well as to establish whether their expression is associated with ischemic atherosclerotic disease. Methods and Results - Human primary T-cells were transduced with lentiviruses encoding distinct FOXP3 isoforms. The phenotype and function of these cells were analyzed by flow cytometry, in vitro suppression assays and RNA-sequencing. We also assessed the effect of activation on Treg cells isolated from healthy volunteers. Treg cell activation resulted in increased FOXP3 expression that predominantly was made up of FOXP3?2. FOXP3?2 induced specific transcription of GARP, which functions by tethering the immunosuppressive cytokine TGF-ß to the cell membrane of activated Treg cells. RT-PCR was used to determine the impact of alternative splicing of FOXP3 in relation with atherosclerotic plaque stability in a cohort of over 150 patients that underwent carotid endarterectomy. Plaque instability was associated with a lower FOXP3?2 transcript usage, when comparing plaques from patients without symptoms and patients with occurrence of recent (<1 month) vascular symptoms including minor stoke, transient ischemic attack or amaurosis fugax. No difference was detected in total levels of FOXP3 mRNA between these two groups. Conclusions - These results suggest that activated Treg cells suppress the atherosclerotic disease process and that FOXP3?2 controls a transcriptional program that acts protectively in human atherosclerotic plaques. Overall design: In this experiment we have analyzed 3 groups of each 3 biological repliactes equalling 9 samples in total.
Publication Title
Alternative Splicing of <i>FOXP3</i> Controls Regulatory T Cell Effector Functions and Is Associated With Human Atherosclerotic Plaque Stability.
Sample Metadata Fields
Subject
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