Project description:ChIP-seq using anti-GFP antibody to map genomic binding of Prdm4-EGFP in stably transfected mouse embryonic stem cells Two independent ChIP-seq replicates for each of two independent Prdm4-EGFP expressing ES cell clones resulting in four anti-GFP ChIP-seq experiments. These samples have paired mouse IgG control.
Project description:ChIP-seq for the strongest cell cycle regulator transcription factors in Drosophila Melanogaster S2 cells. These assays have been used to validate the direct transcriptional targets of the same transcription factors investigated in RNA-seq (E-MTAB-1364) and Affymetrix microarray experiments (E-MTAB-453). ChIP-seq assays have been done with tagged fusion proteins (for example, since we dont have functional E2f antibodies against endogenous E2f , we are transfecting v5-tagged-E2f-ORF to S2 cells and then use antibodies against v5 to detect the signal from E2f binding). If the ChIP-seq has been done with tagged fusion proteins (such as v5-tagged-E2f-ORF), the protein expression has been induced with CuSO4 treatment 48h prior to cell crosslinking & lysis. Our fusion protein constructs are driven by metallothionein promoter, which is induced by CuSO4. E-MTAB-1648, E-MTAB-1364 and E-MTAB-453 are all data from: Bonke M, et al. (2013) Transcriptional networks controlling the cell cycle. G3 (Bethesda) 3, 75-90, PMID: 23316440.
Project description:Characterizing RanBPM interactors in the nucleus and cytoplasm using subcellular fractionation of HeLa cells and affinity purification mass spectrometry.
Project description:The glucocorticoid receptor (GR) regulates gene expression throughout the human genome in a cell-type-specific manner, governing various aspects of homeostasis. The influence of this transcriptional regulator is seen in multiple pathologies, including cancer. Pharmacological activation of the GR is frequently used to alleviate side-effects caused by therapy for patients with various non-lymphoid solid (i.e. non-hematologic) cancers; however, prior studies have shown that this treatment might also have anti-proliferative action on cancer cells. Nonetheless, the molecular underpinnings of glucocorticoid action and its direct effectors in non-lymphoid solid cancers remain elusive. Here, we study the molecular mechanisms of glucocorticoid response in non-lymphoid solid cancers models, focusing on lung cancer. Activation of the GR induces reversible cancer cell dormancy in which cells are tolerant to large array of anti-cancer drugs. This state transition is accompanied by induction of growth factor survival signalling (IGF-1R) and acquired vulnerability to inhibitors of this pathway, both in vitro and in vivo. The observed phenotype is dependent on a single GR target gene - CDKN1C, which encodes for a cell cycle inhibitor protein p57. We have discovered an upstream distal enhancer of CDKN1C, which through GR-induced chromatin looping regulates the expression of this key gene, as confirmed in human tumour specimens. Furthermore, we demonstrate that the SWI/SNF complex composition fine-tunes the GR transcriptional activity at the CDKN1C locus, allowing for precise regulation of cell dormancy induction. Collectively, we show that SWI/SNF-facilitated regulation of CDKN1C underlines GR-induced reversible drug-tolerant state in cancer cells. These insights illustrate the importance of GR signalling in non-lymphoid solid cancer biology, particularly in lung cancer, and warrant caution for use of glucocorticoids in treatment of anti-cancer therapy related side-effects.
Project description:Anterior-posterior differences in H3K27me3 and Ring1B enrichment over the 5 prime Hoxd genes in E10.5 murine distal forelimbs. Chromatin immunoprecipitation (ChIP) of H3K27me3 together with Ring1B and by ChIP-on-chip analysis demonstrated that over the 5 prime HoxD locus H3K27me3 enrichment is decreased and Ring1B enrichment is sparse in limb cells derived from the distal posterior forelimb bud of E10.5 mouse embryos. Array design includes 2 biological replicates for H3K27me3 in the cell lines and Ring1B in the limb tissue, and 2 biological replicates and 2 dye swap replicates for H3K27me3 in the limb tissue.
Project description:The bHLH transcription factor Tfe3 is a powerful regulator of pluripotency and we report a genome-wide analysis of Tfe3 occupancy in mouse ES cells. Nuclear localization of Tfe3 is inhibited by a protein complex containing the tumor-suppressor Folliculin (Flcn) and we also determine Tfe3 binding sites in ES cells expressing an shRNA targeting Flcn. Specificity is controlled for by using unspecific IgGs and ES cells expressing an shRNA targeting Tfe3. ChIP-Seq profiling of Tfe3 in ES cells
Project description:The p52 isoform of Psip1/Ledgf links histone H3K36 methylation and the regulation of alternative splicing. Chromatin immunoprecipitation (ChIP) of Psip1 together with H3K36me3 and Srsf1 and by ChIP-on-chip analysis demonstrated that H3K36me3, Psip1 and SRSF1 enrichment correlates on the gene bodies Array design includes 2 dye swap replicates for Srsf1 and Psip1-/- samples, and single arrays for PSIP and H3K36me3 samples
Project description:The p52 isoform of Psip1/Ledgf links histone H3K36 methylation and the regulation of alternative splicing. Chromatin immunoprecipitation (ChIP) of Psip1 together with H3K36me3 and H3K4me3 and by ChIP-on-chip analysis demonstrated that, Like H3K36me3, Psip1 is enriched on exons of highly expressed genes, Comparision of H3K36me3 and Psip1 binding sites on the expressed and non expressed genes.