Project description:We found acetyl-CoA levels increase when cells are committed to growth. We also found 3 components of the SAGA complex, Spt7p, Sgf73p and Ada3p as well as histones are dynamically acetylated in tune with the acetyl-CoA levels. ChIP-seq study reveals SAGA and H3K9ac predominantly occupy growth genes at the OX growth phase of the yeast metabolic cycle indicating acetyl-CoA levels may drive growth gene transcription program through acetylation of these proteins. Examination of H3K9ac and SAGA binding over two timepoints using H3 and Input as controls
Project description:Under continuous, glucose-limited conditions, budding yeast exhibit robust metabolic cycles associated with major oscillations of gene expression and metabolic state. However, how such fluctuations might be coordinately linked to changes in chromatin status is less well understood. Here, we examine the correlated genome-wide transcription and chromatin states across the yeast metabolic cycle (YMC) at unprecedented temporal resolution, revealing a "just in time supply chain" by which specific cellular processes such as ribosome biogenesis are coordinated in time with remarkable precision. We identify distinct chromatin and splicing patterns associated with different gene categories and determine the relative timing of chromatin modifications to maximal transcription. Additionally, we interrogate chromatin modifier occupancy and observe subtly distinct spatial and temporal patterns compared to the modifications themselves. Furthermore, we identify multiple lysine mutants in H3 or H4 tails that disrupt metabolic cycling, supporting a potentially cooperative role of histone modifications in the YMC. 16 time points RNA-seq and ChIP-seq of 8 histone marks over one metabolic cycle, 14 time points ChIP-seq of 3 chromatin modifiers over one metabolic cycle
Project description:ChIP-seq experiments were performed for the putative telomere repeat-binding factor (PfTRF) in the malaria parasite Plasmodium falciparum strain 3D7. The gene encoding this factor (PF3D7_1209300) was endogenously tagged with either a GFP- or a 3xHA-tag and these transgenic parasite lines were used in ChIP-sequencing experiments. Sequencing of the ChIP and input libraries showed enrichment of PfTRF at all telomere-repeat containing chromosome ends (reference genome Plasmodium falciparum 3D7 from PlasmoDB version 6.1) as well as in all upsB var promoters.In addition,PfTRF was enriched at seven additional, intra-chromosomal sites and called in the PfTRF-HA ChIP-seq only. Plasmodium falciparum 3D7 parasites were generated with -GFP or -3xHA C-terminal tagged TRF (PF3D7_1209300). Nuclei were isolated from formaldehyde cross-linked schizont-stage transgenic parasites and used to prepare chromatin. Chromatin immunoprecipitations were performed using mouse anti-GFP (Roche Diagnostics, #11814460001) or rat anti-HA 3F10 (Roche Diagnostics, #12158167001). Sequencing libraries were prepared according to a Plasmodium-optimized library preparation procedure including KAPA polymerase-mediated PCR amplification.
Project description:Instructive mechanisms are present for induction of DNA methylation, as shown by methylation of specific CpG islands (CGIs) by specific inducers and in specific cancers. However, instructive factors involved are poorly understood, except for involvement of low transcription and trimethylation of histone H3 lysine 27 (H3K27me3). Here, we used methylated DNA immunoprecipitation (MeDIP) combined with a CGI oligonucleotide microarray analysis, and identified 5510 and 521 genes with promoter CGIs resistant and susceptible, respectively, to DNA methylation in prostate cancer cell lines. Expression analysis revealed that the susceptible genes had low transcription in a normal prostatic epithelial cell line. Chromatin immunoprecipitation with microarray hybridization (CHiP-chip) analysis of RNA polymerase II (Pol II) and histone modifications showed that, even among the genes with low transcription, the presence of Pol II was associated with marked resistance to DNA methylation (OR = 0.22; 95% CI = 0.12-0.38), and H3K27me3 was associated with increased susceptibility (OR = 11.20; 95% CI = 7.14-17.55). The same was true in normal human mammary epithelial cells for 5430 and 733 genes resistant and susceptible, respectively, to DNA methylation in breast cancer cell lines. These results showed that the presence of Pol II, active or stalled, and H3K27me3 can predict the epigenetic fate of promoter CGIs independently of transcription levels. To analyze DNA methylation status in normal and cancer cells, MeDIP-CGI oligonucleotide microarray analysis was performed. To analyze expression and histone modification status in normal cells, GeneChip analysis and ChIP-oligonucleotide microarray analysis were performed.
Project description:We performed ChIP-seq targeting the glucocorticoid receptor (GR) in the U2OS-GR cell line. The cell line is derived from U2OS ATTC:HTB-96 and stably transfected with an expression construct for rat GR. The cells were treated with 100 nM dexamethasone for 4 hours, washed 2x with PBS and cultured in hormone-free medium for 24 hours before harvest.
Project description:High-resolution genome-wide mapping of the yeast transcription machinery. ChIP-chip was performed to identify the genomic binding locations for each factor. <br><br>Processed data files are also available on the FTP server for this experiment.
Project description:Expression profiling of Nup153 RNAi-mediated depletion in Drosophila S2 and Kc cells. This experiment is related to experiment E-MEXP-2525.
Project description:ChIP-seq study analysing adult Drosophila melanogaster head, glial, neuronal and fat body, as well as embryonic RNA pol II and H2A.v binding by employing the GAL4-UAS system to generate GFP-fusion proteins and ChIP-seq