We describe a chemical method to label and purify 4-thiouridine (s4U) -containing RNA. We demonstrate that methanethiolsulfonate (MTS) reagents form disulfide bonds with s4U more efficiently than the commonly used HPDP-biotin, leading to higher yields and less biased enrichment. This increase in efficiency allowed us to use s4U-labeling to study global microRNA (miRNA) turnover in proliferating cultured human cells without perturbing global miRNA levels or the miRNA processing machinery. This improved chemistry will enhance methods that depend on tracking different populations of RNA such as 4-thiouridine-tagging to study tissue-specific transcription and dynamic transcriptome analysis (DTA) to study RNA turnover. Overall design: s4U metabolic labeling of RNA in 293T cells, followed by biochemical enrichment of labeled RNA with two biotinylation reagents, RNAs >200nt and miRNAs in separate experiments
Tracking Distinct RNA Populations Using Efficient and Reversible Covalent Chemistry.
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View SamplesThis SuperSeries is composed of the SubSeries listed below.
Human LSD2/KDM1b/AOF1 regulates gene transcription by modulating intragenic H3K4me2 methylation.
Cell line
View SamplesDynamic histone H3K4 methylation is an important epigenetic component of transcriptional regulation. However, most of our current understanding of this histone mark is confined to regulation of transcriptional initiation. We now show that human LSD2/KDM1b/AOF1, the human homolog of LSD1, is a novel H3K4me1/2 demethylase that specifically regulates histone H3K4 methylation within intragenic regions of its target genes. Genome-wide mapping reveals that LSD2 associates predominantly with the gene bodies of actively transcribed genes, but is markedly absent from promoters. Depletion of endogenous LSD2 results in an increase of H3K4me2 as well as a decrease of H3K9me2 at LSD2 binding sites, and a consequent dysregulation of target gene transcription. Furthermore, characterization of LSD2 complex revealed that LSD2 forms active complexes with euchromatic histone methyltransferases EHMT1/2 and NSD3 as well as cellular factors involved in active transcription elongation. These data provide a possible molecular mechanism linking LSD2 to transcriptional regulation post initiation.
Human LSD2/KDM1b/AOF1 regulates gene transcription by modulating intragenic H3K4me2 methylation.
Cell line
View SamplesLipotoxicity is a metabolic disorder that results from accumulation of lipids, particularly fatty acids, in non-adipose tissue leading to cellular dysfunction, lipid droplet formation, activate the ATF3 stress pathway, induce secretion of inflammatory cytokines, and increase APP. Our observations are consistent with neurovascular lipotoxicity that could play a role in cognitive decline with aging.
Triglyceride-rich lipoprotein lipolysis products increase blood-brain barrier transfer coefficient and induce astrocyte lipid droplets and cell stress.
Specimen part, Treatment
View SamplesTo identify the gene expression profile of enteric glia and assess the transcriptional similarity between enteric and extraenteric glia, we performed RNA sequencing analysis on PLP1-expressing cells in the mouse intestine. This analysis shows that enteric glia are transcriptionally unique and distinct from other cell types in the nervous system. Enteric glia express many genes characteristic of the myelinating glia, Schwann cells and oli- godendrocytes, although there is no evidence of myelination in the murine ENS. Overall design: Total RNA expression profiles of PLP1 expressing enteric glial cells (GFP+) and non-glial cells (GFP-negative) were obtained from the ileum and colon of juvenile PLP1-eGFP transgenic mice.
Enteric glia express proteolipid protein 1 and are a transcriptionally unique population of glia in the mammalian nervous system.
Specimen part, Cell line, Subject
View SamplesEffects of SOCS3 on the transcriptional response of bone marrow-derived macrophages to IL-6.
SOCS3 regulates the plasticity of gp130 signaling.
No sample metadata fields
View SamplesEpigenetic changes play a role in the pathogenesis of myeloid malignancies and hypomethylating agents have shown efficacy in these diseases. We studied the apoptotic effect, the genome-wide methylation and gene expression profiles in HL60 cells following decitabine treatment, using micro-array technologies. Decitabine treatment resulted in a decrease in global DNA methylation, corresponding to 4876 probeset IDs with significantly reduced methylation levels, while expression of 2583 IDs was induced. The integrated analysis identified 160 genes demethylated and upregulated by decitabine, mainly including development and differentiation pathways genes. Genes target of polycomb group protein regulation were overrepresented in this group. Apoptosis was induced by decitabine and apoptosis-specific PCR arrays more precisely indicated decitabine-induced upregulation of 13 apoptosis-related genes, in particular Dap-kinase 1 and Bcl2L10. Correspondingly, in primary patient samples, BCL2L10 was hypermethylated in 45% of AML, 43% of therapy-related myeloid neoplasms, 12% of MDS and in none of the controls.
Analysis of genome-wide methylation and gene expression induced by 5-aza-2'-deoxycytidine identifies BCL2L10 as a frequent methylation target in acute myeloid leukemia.
Specimen part
View SamplesElimination of peripheral retinal axons leads to changes in gene expression in both visual and somatosensory thalamic neurons.
Prenatal thalamic waves regulate cortical area size prior to sensory processing.
Specimen part
View SamplesBiochemical fractionation of HEK293 nuclei and RNA-seq of chromatin-associated and soluble-nuclear RNA. Overall design: Nuclei from three biological replicates were isolated by detergent lysis, fractionated, then three chromatin and three soluble RNA samples were converted to cDNA using Illumina TruSeq stranded protocol, and sequenced on Illumina HiSeq2000
Nuclear Fractionation Reveals Thousands of Chromatin-Tethered Noncoding RNAs Adjacent to Active Genes.
No sample metadata fields
View SamplesMorphogenesis of cellecting duct system within developing mouse kidney is driven by growth at the tips of ureteric epithelium. To characterize the transcription program within the tip compartment, here we performed mRNA-Seq of tip cells (Wnt11RFP+;Hoxb7+ cells) and stalk cells (Wnt11RFP-;Hoxb7GFP+ cells) obtained from mouse embryonic kidney through FACS. We identified tip-specific genes from these data, and verified with in situ hybridization and followed up with mechanistic study for some of the intersting targets. Overall design: Examination of two cell types within the ureteric bud of the developing mouse kidney
Cellular heterogeneity in the ureteric progenitor niche and distinct profiles of branching morphogenesis in organ development.
Cell line, Subject
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