To determine whether immortalized cells derived from the rat SCN (SCN 2.2) retain intrinsic rhythm-generating properties characteristic of the SCN, oscillatory properties of the SCN2.2 transcriptome were analyzed and compared to those found in the rat SCN in vivo using rat U34A Affymetrix GeneChips. SCN2.2 cells were expanded in 6-well plates. At 6-hour interval across 2 circadian cycles, cells from single 6-well plates were harvested and pooled for total RNA extraction.
Circadian profiling of the transcriptome in immortalized rat SCN cells.
No sample metadata fields
View SamplesTo screen for specific circadian outputs that may distinguish the pacemaker in the mammalian suprachiasmatic nucleus (SCN) from peripheral-type oscillators in which the canonical clockworks are similarly regulated in a circadian manner, the rhythmic behavior of the transcriptome in forskolin-stimulated NIH/3T3 fibroblasts was analyzed and compared to that found in the rat SCN in vivo and SCN2.2 cells in vitro. Similar to the scope of circadian gene expression in SCN2.2 cells and the rat SCN, NIH/3T3 fibroblasts exhibited circadian fluctuations in the expression of the core clock genes, Per2, Bmal1 (Mop3), and Cry1 and 323 functionally diverse transcripts (2.6%), many of which were involved in cell communication. Overlap in rhythmically-expressed transcripts among NIH/3T3 fibroblasts, SCN2.2 cells and the rat SCN was limited to these clock genes and four other genes that mediate fatty acid and lipid metabolism or function as nuclear factors. Compared to NIH/3T3 cells, circadian gene expression in SCN oscillators was more prevalent among cellular pathways mediating glucose metabolism and neurotransmission. Coupled with evidence for the rhythmic regulation of the inducible isoform of nitric oxide synthase, the enzyme responsible for the production of nitric oxide, in SCN2.2 cells and the rat SCN but not in fibroblasts, studies examining the effects of a NOS inhibitor on metabolic rhythms in co-cultures containing SCN2.2 cells and untreated NIH/3T3 cells suggest that this gaseous neurotransmitter may play a key role in SCN pacemaker function. Thus, this comparative analysis of circadian gene expression in SCN and non-SCN cells may have important implications in the selective identification of circadian signals involved in the coupling of SCN oscillators and the regulation of rhythmicity in downstream cells or tissues.
Circadian profiling of the transcriptome in NIH/3T3 fibroblasts: comparison with rhythmic gene expression in SCN2.2 cells and the rat SCN.
No sample metadata fields
View SamplesNovel strategies are needed to modulate -cell differentiation and function as potential -cell replacement or restorative therapies for diabetes. We previously demonstrated that small molecules based on the isoxazole scaffold drive neuroendocrine phenotypes. The nature of the effects of isoxazole compounds on cells was incompletely defined. We find that isoxazole largely induced genes that support neuroendocrine and -cell phenotypes, and suppressed a set of genes important for proliferation. Isoxazole alters -cell metabolites and protects glucose-responsive signaling pathways under lipotoxic conditions. Finally, we show that isoxazole improves glycemia in a mouse model of -cell regeneration. Isoxazole is a prime candidate for altering cell fate in different contexts.
Isoxazole Alters Metabolites and Gene Expression, Decreasing Proliferation and Promoting a Neuroendocrine Phenotype in β-Cells.
Specimen part
View SamplesBackground: Weaning of beef calves is a necessary husbandry practice and involves separating the calf from its mother, resulting in numerous stressful events including dietary change, social reorganisation and the cessation of the maternal-offspring bond and is often accompanied by housing. While much recent research has focused on the physiological response of the bovine immune system to stress in recent years, little is known about the molecular mechanisms modulating the immune response. Therefore, the objective of this study was to provide new insights into the molecular mechanisms underlying the physiological response to weaning at housing in beef calves using Illumina RNA-seq. Results: The leukocyte transcriptome was significantly altered for at least 7 days following either housing or weaning at housing. Analysis of differentially expressed genes revealed that four main pathways, cytokine signalling, transmembrane transport, haemostasis and G-protein-coupled receptor (GPRC) signalling, were differentially regulated between control and weaned calves and underwent significant transcriptomic alterations in response to weaning stress on day 1, 2 and 7. Of particular note, chemokines, cytokines and integrins were consistently found to be up-regulated on each day following weaning. Evidence for alternative splicing of genes was also detected, indicating that a number of genes involved in the innate and adaptive immune response may be alternatively transcribed, including those responsible for toll receptor cascades and T cell receptor signalling. Conclusions: This study represents the first application of RNA-Seq technology for genomic studies in bovine leukocytes in response to weaning stress. Weaning stress induces the activation of a number of cytokine, chemokine and integrin transcripts and may alter the immune system whereby the ability of a number of cells of the innate and adaptive immune system to locate and destroy pathogens is transcriptionally enhanced. Stress alters the homeostasis of the transcriptomic environment of leukocytes for at least 7 days following weaning, indicating long-term effects of stress exposure in the bovine. The identification of gene signature networks that are stress activated provides a mechanistic framework to characterise the multifaceted nature of weaning stress adaptation in beef calves. Thus, capturing subtle transcriptomic changes provides insight into the molecular mechanisms that underlie the physiological response to weaning stress. Overall design: Examination of a time course (day 0, 1, 2 and 7) for 2 treatments, calves either housed with their dam (control) or housed and simultaneously weaned, using RNA-seq. The supplementary processed data file 'read_counts.txt' contains unnormalized read counts for each Ensembl bovine gene in each of the 48 samples. Unnormalized counts are required for input to EdgeR. Genome build: Btau4.0
Transcriptomic analysis of the stress response to weaning at housing in bovine leukocytes using RNA-seq technology.
Specimen part, Disease, Treatment, Subject
View SamplesThe purpose of this study was to profile gene expression changes that could occur with the loss of co-repressor SLIRP. In addition, we wanted to investigate how DHT could further alter gene expression. Methods: LNCaP cells were transfected with custom nonsense control siRNA or a pool of SLIRP siRNA (HSS130109, HSS188666, HSS188667 Invitrogen, Carlsbad, CA) at 40nM for 24hrs. Cells were washed once with PBS and replaced with SFM containing EtOH or 1nM/ml DHT for another 24hrs. Two biological replicates were collected from 2 different experiemnts for total of 4 replicates. RNA collected was sequenced using Illumia HiSeq2000 single read 50 bp by the HTSF core at UNC and aligned and normalized by the Bioinformatics core at UNC. Results: We found a differential gene expression pattern between our control and SLIRP knockdown samples. We also identified a 176 AR gene signature with 3 subclasses and a large SLIRP gene signature (~1700). Overall design: Examination of control (NS) vs. SLIRP siRNA treated with 1nM DHT in the prostate cancer cell line LNCaP
Interaction between androgen receptor and coregulator SLIRP is regulated by Ack1 tyrosine kinase and androgen.
No sample metadata fields
View SamplesWe performed RNA sequencing on melanopsin deleted retinas (Opn4-DTA/DTA) to determine potential cues involved in instructing cone photoreceptor positioning Overall design: RNAseq of whole P8 retinal extracts from wild-type littermate vs. Opn4DTA/DTA mice
Melanopsin Retinal Ganglion Cells Regulate Cone Photoreceptor Lamination in the Mouse Retina.
Specimen part, Subject
View SamplesIncreased energy demands to support lactation, coupled with lowered feed intake capacity results in negative energy balance (NEB) and is typically characterized by extensive mobilization of body energy reserves in the early postpartum dairy cow. The catabolism of stored lipid leads to an increase in the systemic concentrations of nonesterified fatty acids (NEFA) and -hydroxy butyrate (BHB). Oxidation of NEFA in the liver result in the increased production of reactive oxygen species and the onset of oxidative stress and can lead to disruption of normal metabolism and physiology. The immune system is depressed in the peripartum period and early lactation and dairy cows are therefore more vulnerable to bacterial infections causing mastitis and or endometritis at this time. A bovine Affymetrix oligonucleotide array was used to determine global gene expression in the spleen of dairy cows in the early postpartum period. Spleen tissue was removed post mortem from five severe NEB (SNEB) and five medium NEB (MNEB) cows 15 days postpartum.SNEB increased systemic concentrations of NEFA and BHB, and white blood cell and lymphocyte numbers were decreased in SNEB animals. A total of 545 genes were altered by SNEB. Network analysis using Ingenuity Pathway Analysis revealed that SNEB was associated with NRF2-mediated oxidative stress, mitochondrial dysfunction, endoplasmic reticulum stress, natural killer cell signaling, p53 signaling, downregulation of IL-15, BCL-2, and IFN- ; upregulation of BAX and CHOP and increased apoptosis with a potential negative impact on innate and adaptive immunity.
Pleiotropic effects of negative energy balance in the postpartum dairy cow on splenic gene expression: repercussions for innate and adaptive immunity.
No sample metadata fields
View SamplesTo determine the role of Mbd3/NuRD in lymphopoiesis, gene expression in purified populations of Mbd3-deleted and control lymphoid progenitor cells was analysed using RNA-seq. Overall design: Mbd3-deficient and control lymphoid progenitors were isolated from mouse bone marrow by flow cytometry, including haematopoietic stem cells (HSCs), lymphoid-primed multipotent progenitors (LMPPs), all-lymphoid progenitors (ALPs) and B cell-biased lymphoid progenitors (BLPs). RNA-seq was performed on 100 HSCs or 150 cells from the other populuations, using the previously described smartseq2 protocol for RNA-seq of small numbers of cells (Picelli et al. (2014) Nature protocols 9:171).
Mbd3/NuRD controls lymphoid cell fate and inhibits tumorigenesis by repressing a B cell transcriptional program.
Sex, Age, Specimen part, Cell line, Subject
View SamplesThis SuperSeries is composed of the SubSeries listed below.
The UBC-40 Urothelial Bladder Cancer cell line index: a genomic resource for functional studies.
Specimen part, Cell line
View SamplesThis is a comprehensive genomic characterization of 40 urothelial bladder carcinoma (UBC) cell lines including information on origin, mutation status of genes implicated in bladder cancer (FGFR3, PIK3CA, TP53, and RAS), copy number alterations assessed using high density SNP arrays, uniparental disomy (UPD) events, and gene expression. Based on gene mutation patterns and genomic changes we identify lines representative of the FGFR3-driven tumor pathway and of the TP53/RB tumor suppressor-driven pathway. High-density array copy number analysis identified significant focal gains (1q32, 5p13.1-12, 7q11, and 7q33) and losses (i.e. 6p22.1) in regions altered in tumors but not previously described as affected in bladder cell lines. We also identify new evidence for frequent regions of UPD, often coinciding with regions reported to be lost in tumors. Previously undescribed chromosome X losses found in UBC lines also point to potential tumor suppressor genes. Cell lines representative of the FGFR3-driven pathway showed a lower number of UPD events. Overall, there is a predominance of more aggressive tumor subtypes among the cell lines. We provide a cell line classification that establishes their relatedness to the major molecularly-defined bladder tumor subtypes. The compiled information should serve as a useful reference to the bladder cancer research community and should help to select cell lines appropriate for the functional analysis of bladder cancer genes, for example those being identified through massive parallel sequencing.
The UBC-40 Urothelial Bladder Cancer cell line index: a genomic resource for functional studies.
Specimen part, Cell line
View Samples