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SRP032276
Saccharomyces cerevisiae
56 Downloadable Samples
Illumina HiSeq 2500, Illumina MiSeq, Illumina HiSeq 2000
Description
N6-methyladenosine (m6A) is the most ubiquitous mRNA base modification, but little is known about its precise location, temporal dynamics, and regulation. Here, we generated genomic maps of m6A sites in meiotic yeast transcripts at nearly single-nucleotide resolution, identifying 1,308 putatively methylated sites within 1,183 transcripts. We validated 8/8 methylation sites in different genes with direct genetic analysis, demonstrated that methylated sites are significantly conserved in a related species, and built a model that predicts methylated sites directly from sequence. Sites vary in their methylation profiles along a dense meiotic time-course, and are regulated both locally, via predictable methylatability of each site, and globally, through the core meiotic circuitry. The methyltransferase complex components localize to the yeast nucleolus, and this localization is essential for mRNA methylation. Our data illuminates a conserved, dynamically regulated methylation program in yeast meiosis, and provides an important resource for studying the function of this epitranscriptomic modification. Overall design: Examination of m6A methylation under various conditions
Publication Title
High-resolution mapping reveals a conserved, widespread, dynamic mRNA methylation program in yeast meiosis.
Sample Metadata Fields
Cell line, Subject
View Samples- Mus musculus
- 16 Downloadable Samples
Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Transfection of dsDNA into many mammalian cell types indues the production of type I interferons and interferon-stimulated genes. We performed an siRNA screen to identify genes involved in this innate immune response, and identified Abcf1.
Publication Title
Identification of regulators of the innate immune response to cytosolic DNA and retroviral infection by an integrative approach.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 2 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
We used microarrays to determine which genes are upregulated by IFNbeta stimulation in 293T cells.
Publication Title
Identification of regulators of the innate immune response to cytosolic DNA and retroviral infection by an integrative approach.
Sample Metadata Fields
Specimen part, Cell line, Treatment
View Samples- Mus musculus
- 656 Downloadable Samples
- IlluminaHiSeq2500
Description
We introduced genome-wide pooled CRISPR-Cas9 libraries into primary mouse dendritic cells (DCs) to identify genes that control the induction of tumor necrosis factor (TNF) by bacterial lipopolysaccharide (LPS), a key process in the host response to pathogens, mediated by the TLR4 pathway. We found many of the known regulators of TLR4 signaling, as well as dozens of previously unknown candidates that we validated. Overall design: We used stain base phenotype (staining for TNF) in order to search for negative and positive regulators of LPS response in differentiated BMDCs
Publication Title
A Genome-wide CRISPR Screen in Primary Immune Cells to Dissect Regulatory Networks.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 28 Downloadable Samples
- Illumina HiSeq 2500
Description
Protein expression is regulated by production and degradation of mRNAs and proteins, but their specific relationships remain unknown. We combine measurements of protein production and degradation and mRNA dynamics to build a quantitative genomic model of the differential regulation of gene expression in LPS stimulated mouse dendritic cells. Changes in mRNA abundance play a dominant role in determining most dynamic fold changes in protein levels. Conversely, the preexisting proteome of proteins performing basic cellular functions is remodeled primarily through changes in protein production or degradation, accounting for over half of the absolute change in protein molecules in the cell. Thus, the proteome is regulated by transcriptional induction of novel cellular functions and remodeling of preexisting functions through the protein life cycle. Overall design: Mouse primary dendritic cells were treated with LPS or mock stimulus and profiled over a 12-hour time course. Cells were grown in M-labeled SILAC media, which was replaced with H-labeled SILAC media at time 0. Aliquots were taken at 0, 0.5, 1, 2, 3, 4, 5, 6, 9, and 12 hours post-stimulation and added to equal volumes of a master mix of unlabeled (L) cells for the purpose of normalization. RNA-Seq was performed at 0, 1, 2, 4, 6, 9, and 12 hours post-stimulation.
Publication Title
Immunogenetics. Dynamic profiling of the protein life cycle in response to pathogens.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 46 Downloadable Samples
- IlluminaHiSeq2500, IlluminaMiSeq
Description
N6-methyladenosine (m6A) is a common modification of mRNA, with potential roles in fine-tuning the RNA life cycle, but little is known about the pathways regulating this process and its physiological role. Here, we used mass-spectrometry to identify a dense network of proteins physically interacting with METTL3, a core component of the methyltransferase complex, and show that two of them, WTAP and KIAA1429, are required for methylation. Combining high resolution m6A-Seq with knockdown of WTAP allowed us to define accurate maps, at near single-nucleotide resolution, of sites of mRNA methylation across four dynamic programs in human and mouse, including development, differentiation, reprogramming and immune response. Internal WTAP-dependent methylation sites were largely static across the different surveyed conditions and present in the majority of mRNAs. However, methylations were found at much lower levels within highly expressed mRNAs, and methylation is inversely correlated with mRNA stability, consistent with a role in establishing an overall basal, cell-type invariant, distribution of degradation rates. In addition, we identify thousands of WTAP-independent methylation sites at transcription initiation sites, forming part of the mRNA cap structure. We show that the methylations occur at the first transcribed nucleotide, and find that thousands of transcripts are present in different isoforms differing in the methylation state of the first transcribed nucleotide, a previously unappreciated complexity of the transcriptome. Together, our data sheds new light on the proteomic and transcriptional underpinnings of this epitranscriptomic modification in mammals. Overall design: Examination of m6A methylation in human Hek293 and A549 cell lines, in human embryonic stem cells (ESCs) undergoing differentiation to neural progenitor cells (NPCs), in OKMS inducible fibroblasts reprogrammed into iPSC, and upon knockdown of factors using siRNAs or shRNAs.
Publication Title
Perturbation of m6A writers reveals two distinct classes of mRNA methylation at internal and 5' sites.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 40 Downloadable Samples
- Illumina HiSeq 2500, Illumina HiSeq 2000
Description
N6-methyladenosine (m6A) is a common modification of mRNA, with potential roles in fine-tuning the RNA life cycle, but little is known about the pathways regulating this process and its physiological role. Here, we used mass-spectrometry to identify a dense network of proteins physically interacting with METTL3, a core component of the methyltransferase complex, and show that two of them, WTAP and KIAA1429, are required for methylation. Combining high resolution m6A-Seq with knockdown of WTAP allowed us to define accurate maps, at near single-nucleotide resolution, of sites of mRNA methylation across four dynamic programs in human and mouse, including development, differentiation, reprogramming and immune response. Internal WTAP-dependent methylation sites were largely static across the different surveyed conditions and present in the majority of mRNAs. However, methylations were found at much lower levels within highly expressed mRNAs, and methylation is inversely correlated with mRNA stability, consistent with a role in establishing an overall basal, cell-type invariant, distribution of degradation rates. In addition, we identify thousands of WTAP-independent methylation sites at transcription initiation sites, forming part of the mRNA cap structure. We show that the methylations occur at the first transcribed nucleotide, and find that thousands of transcripts are present in different isoforms differing in the methylation state of the first transcribed nucleotide, a previously unappreciated complexity of the transcriptome. Together, our data sheds new light on the proteomic and transcriptional underpinnings of this epitranscriptomic modification in mammals. Overall design: Examination of m6A methylation across different knockdowns using shRNAs in mouse embryonic fibroblasts, in embyronic and adult brains, and in dendritic cell stimulated with LPS.
Publication Title
Perturbation of m6A writers reveals two distinct classes of mRNA methylation at internal and 5' sites.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 4 Downloadable Samples
- Affymetrix HT Mouse Genome 430A Array (htmg430a)
Description
We used microarrays to detail the global programme of gene expression underlying Polo-like kinase inhibition with BI 2536 compound in mouse bone marrow-derived dendritic cells (BMDCs) stimulated with Toll-like receptor agonists LPS and poly(I:C).
Publication Title
Systematic discovery of TLR signaling components delineates viral-sensing circuits.
Sample Metadata Fields
Specimen part
View Samples- Caenorhabditis elegans
- 2 Downloadable Samples
- Illumina HiSeq 2000
Description
PMK-1 is involved in the heat stress response of C. elegans, translocates to the nucleus upon heat exposure and influences the expression of chaperone genes, proteasomal subunits and protein-biosynthesis related genes. Overall design: Differential Gene expression of WT and pmk-1 deletion mutant (KU25) after 5 hours at 35°C
Publication Title
The p38 MAPK PMK-1 shows heat-induced nuclear translocation, supports chaperone expression, and affects the heat tolerance of Caenorhabditis elegans.
Sample Metadata Fields
Cell line, Subject
View Samples- Mus musculus
- 21 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Complex regulatory mechanisms control continuous maintenance of myeloid progenitors and renewal of differentiated cells. Transcription factors play a important role in these processes. Here we report that the activation the calcineurin-NFAT signaling pathway inhibit the proliferation of myeloid granulocyte-monocyte progenitor (GMP). Myeloid progenitor subtypes possessed different susceptibilities to Ca2+ flux induction and consequently differential engagement of the calcineurin-NFAT pathway. This study show that inhibition of the calcineurin-NFAT pathway enhanced proliferation of GMPs both in vivo and in vitro. The calcineurin-NFAT signaling in GMPs is initiated through Flt3-L. The inhibition of the calcineurin-NFAT pathway altered the expression of the cell cycle regulation genes CDK4, CDK6, and CDKN1A, thus enabling faster cell cycle progression. The extensive use of NFAT inhibitors in the clinic should take into account that, in addition to the immunosuppression role in lymphoid cells, these NFAT inhibitors also affect the maintenance of the myeloid compartment.
Publication Title
Calcium and calcineurin-NFAT signaling regulate granulocyte-monocyte progenitor cell cycle via Flt3-L.
Sample Metadata Fields
Specimen part
View Samples