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SRP132308
Drosophila melanogaster
12 Downloadable Samples
Illumina HiSeq 2000
Description
Purpose: Investigating the role of Drosophila G9a in oxidative stress responses. Methods: Flies were collected after eclosion and allowed to recover from CO2 exposure for 5 days prior to paraquat exposure. Paraquat (Methyl viologen dichloride hydrate 98 %; Sigma 856177) was mixed into the flyfood at 40 °C to a final concentration of 50 mM. For OS induction, 5-9 day old flies were transferred to paraquat containing food and incubated at 25 °C and 70 % humidity. At each time point, flies were flash frozen in liquid nitrogen followed by vortexing and filtering through a series of sieves to isolate heads from other body parts. 200 fly heads per sample were used for RNA extraction using QIAGEN lipid mini tissue kit. The TruSeq RNA Sample Preparation Kit v2 (Illumina) was used to prepare adapter ligated PCR fragments for sequencing. PCR was used to selectively enrich the fragments containing the adapters. The PCR fragments were validated using Agilent 2200 TapeStation. Single indexed samples were multiplexed and sequenced on an Illumina HiSeq 2000 sequencing system (Illumina) in single-end mode with a read length of 35 bp. Quality of sequenced reads was assessed with FastQC. The RNAseq experiments were conducted on two biological duplicates for each condition. Sequenced reads were aligned with Burrows-Wheeler algorithm (BWA) (Li & Durbin, 2010) to the Drosophila reference genome (BDGP.5, http://www.fruitfly.org/) and per gene read counts were generated with HTSeq count (Anders et al, 2015). 25–30 million reads with high quality alignment were obtained for each sample and used for differential expression analysis. DESeq (Anders & Huber, 2010) was used to obtain library size-normalized read counts and to calculate differential expression of genes in 4 pairwise comparisons: 0 h versus 6 h and 0 h versus 12 h after OS in both G9a mutants and controls (fold change =1.5, adjusted p-value= 0.05, Benjamini-Hochberg). Results: We found 2731 genes to be differential expressed in at least one of the four pairwise comparisons. The largest group of differentially expressed (DE) genes are highly augmented upon OS induction in the G9a mutant (41.7 % of all DE genes). The second largest group of DE genes (23.9 % of all DE genes) were more downregulated in G9a mutant in response to OS. Genes that are over-activated in G9a mutants are predominantly involved in OS response and OS mediated damage, whereas genes that are downregulated in G9a mutants are involved in energy metabolism. Conclusions: Our data suggest that G9a provides an epigenetic mechanism that safeguards an appropriate transcriptional response to OS and preserves immediately available energy, thereby acting as a critical regulatory hub between the transcriptional and physiological responses to oxidative stress. Overall design: fly-head mRNA libraries of 5-9 days old male G9aDD1 mutant and control during 0, 6 and 12 hours of paraquat oxidative stress exposure were sequenced in duplicate on Hi-seq 2000.
Publication Title
The histone methyltransferase G9a regulates tolerance to oxidative stress-induced energy consumption.
Sample Metadata Fields
Specimen part, Treatment, Subject
View Samples- Homo sapiens
- 24 Downloadable Samples
- IlluminaHiSeq2000
Description
FOXE3 is a lens specific transcription factor that has been associated with anterior segment ocular dysgenesis. To determine the transcriptional target(s) of FOXE3 that are indispensable for the anterior segment development, we examined the transcriptome and the proteome of cells expressing truncated FOXE3 responsible for Peters anomaly identified through linkage-coupled next-generation whole exome sequencing. We found that DNAJB1, an autophagy-associated protein, was the only candidate exhibiting differential expression in both screens. We confirmed the candidacy of DNAJB1 through chromatin immunoprecipitation and luciferase assays while knockdown of DNAJB1 in human lens epithelial cells resulted in mitotic arrest. Subsequently, we targeted dnajb1a in zebrafish through injection of a splice-blocking morpholino. The dnajb1a morphants exhibited underdeveloped cataractous lenses with persistent apoptotic nuclei. In conclusion, we have identified DNAJB1 as a transcriptional target of FOXE3 in a novel pathway that is crucial for development of the anterior segment of the eye. Overall design: Human Embryonic Kidney (HEK293FT) cells were transfected with the expression vector (pT-RexTM-DEST30) harboring either the wild type or the mutant (C240*) FOXE3 ORF (open reading frame). The experimental design included a total of eight biological replicates of cells expressing the wild type and eight replicates of mutant FOXE3 along with eight non-transfected controls. Cells were harvested 24-hour post-transfection and subjected to total RNA isolation for the preparation of whole transcriptome next-generation sequencing libraries. Initially, we examined the quality of transcriptome libraries on a MiSeq genome analyzer. Subsequent to confirmation of the quality, all libraries were paired-end sequenced (2 x 100 bp) using Illumina TruSeq Cluster V3 flow cell at a concentration of 13.0 pM in two separate lanes (12 bar-coded mRNA pooled libraries in each lane) on a HiSeq 2000 genome analyzer.
Publication Title
FOXE3 contributes to Peters anomaly through transcriptional regulation of an autophagy-associated protein termed DNAJB1.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 20 Downloadable Samples
Affymetrix Human Exon 1.0 ST Array [probe set (exon) version (huex10st)
Description
Gene expression analysis under normal culture conditions (RPMI-10%FBS) and at optimal cell densities.
Publication Title
Low-risk susceptibility alleles in 40 human breast cancer cell lines.
Sample Metadata Fields
Cell line
View Samples- Mus musculus
- 12 Downloadable Samples
Description
Purpose: Transcriptome is the entire repertoire of all transcripts present in a cell at any particular time. We undertook next-generation whole transcriptome sequencing approach to gain insight of the transcriptional landscape of the developing mouse lens. Methods: We ascertained mice lenses at six developmental time points including two embryonic (E15 and E18) and four postnatal stages (P0, P3, P6, and P9). The ocular tissue at each time point was maintained as two distinct pools serving as biological replicates for each developmental stage. The mRNA and small RNA libraries were paired-end sequenced on Illumina HiSeq 2000 and subsequently analyzed using bioinformatics tools. Results: Mapping of mRNA and small RNA libraries generated 187.56 and 154.22 million paired-end reads, respectively. We detected a total of 14,465 genes in the mouse ocular lens. Of these, 46 genes exhibited 40-fold differential expression compared to transcriptional levels at E15. Likewise, small RNA profiling identified 379 microRNAs (miRNAs) expressed in mouse lens. Of these, 49 miRNAs manifested an 8-fold or higher differential expression when compared, as above to the microRNA expression at E15. Conclusion: We report the first comprehensive profile of developing murine lens transcriptome including both mRNA and miRNA through next-generation RNA sequencing. A complete repository of the lens transcriptome of six developmental time points will be monumental in elucidating processes essential for development of the ocular lens and maintenance its transparency. Overall design: Whole transcrtiome and microRNA profilling of mouse lens using 2 embryonic (E15 and E18) and 4 postnatal stages (P0, P3, P6 and P9) in duplicates through high-throughput sequening using Illumina HiSeq2000.
Publication Title
Identification of novel transcripts and peptides in developing murine lens.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 12 Downloadable Samples
- Illumina HiSeq 2500
Description
Comparing WT mice to a mouse model of mental retardation, this work identifies genes which display differences in ribosome-bound mRNAs, in hippocampus CA1 pyramidal cells. These genes products are potent functional components of neuronal plasticity and hippocampus-dependent memory. Overall design: Using a triple transgenic mouse line, we immunoprecipitated the HA-Rpl22 protein to isolate and sequence ribosome-associated mRNA in CA1 pyramidal cells. Pairwise comparison of wild type and Fmr1 KO mice defined a specific gene expression profile.
Publication Title
Cell Type-Specific mRNA Dysregulation in Hippocampal CA1 Pyramidal Neurons of the Fragile X Syndrome Mouse Model.
Sample Metadata Fields
Specimen part, Subject
View Samples- Homo sapiens
- 3 Downloadable Samples
- Affymetrix Human Gene 2.0 ST Array (hugene20st)
Description
Transcriptome analysis on ING5-knockdown brain tumor stem cell lines
Publication Title
ING5 activity in self-renewal of glioblastoma stem cells via calcium and follicle stimulating hormone pathways.
Sample Metadata Fields
Specimen part, Cell line
View Samples- Homo sapiens
- 9 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
Many studies have characterized the results of shear stress changes on cultured endothelial cells in different bioreactor systems. However it is still unclear how an invasive intervention like stent procedure may influence the transcriptional response of endothelium.
Publication Title
Vascular injury post stent implantation: different gene expression modulation in human umbilical vein endothelial cells (HUVECs) model.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 8 Downloadable Samples
- Illumina HiSeq 2500
Description
In order to determine the imprinted transcription factor Zac1 targets, we overexpressed Zac1 in a mouse insulinoma cell line and measured the regulated expressed genes by RNA-seq. We have shown that Zac1 regulates many genes belonging to the Imprinted Gene Network, including genes coding for the extra-cellular matrix. Overall design: Determination of Zac1 target genes in transfected Min6 cells (4 biological replicates) using RNA-seq, .
Publication Title
Identification of Plagl1/Zac1 binding sites and target genes establishes its role in the regulation of extracellular matrix genes and the imprinted gene network.
Sample Metadata Fields
Specimen part, Subject
View Samples- Homo sapiens
- 18 Downloadable Samples
- IlluminaHiSeq2000
Description
RNAPII pausing/termination shortly after initiation is a hallmark of gene regulation. However, the molecular mechanisms involved are still to be uncovered. Here, we show that NELF interacts with Integrator complex subunits (INTScom) forming a stable complex with RNPII and Spt5. The interaction between NELF and INTScom subunits is RNA and DNA independent. Using both HIV-1 promoter and genome wide analyses, we demonstrate that Integrator subunits specifically control NELF-mediated RNAPII pause/release at coding genes. The strength of RNAPII pausing is determined by the nature of the NELF-associated complex. Interestingly, in addition to controlling RNAPII pause release INTS11 catalytic subunit of the INTScom is required for the synthesis of full length mRNA. Finally, INTScom-target genes are enriched in HIV-1 TAR/ NELF-binding element and in a 3'box sequence required for snRNA biogenesis. Revealing these unexpected functions of INTScom in regulating RNAPII pausing/release and completion of mRNA synthesis of NELF-target genes will contribute to our understanding of the gene expression cycle. Overall design: Genome-wide expression in HeLa cells in the absence of Integrator 11, or NELF or mock (control) depleted by strand-specific RNASeq (Illumina)
Publication Title
Integrator complex regulates NELF-mediated RNA polymerase II pause/release and processivity at coding genes.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 9 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Transcription factor FoxQ1 belongs to family of Fox transcription factors containing forkhead box (winged helix) domain. This family of proteins plays an important role in cell growth, proliferation, and differentiation. Expression of FoxQ1 was found in cancer cells and was associated with increased migration and invasion. Recently, elevated FoxQ1 expression was found in IL4 stimulated macrophages.
Publication Title
IL-4 driven transcription factor FoxQ1 is expressed by monocytes in atopic dermatitis and stimulates monocyte migration.
Sample Metadata Fields
Cell line
View Samples