Genome wide profiling of RNA polymerase III in Saccharomyces cerevisiae
Ontology highlight
ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE39566 | GEO | 2013/06/21
SECONDARY ACCESSION(S): PRJNA171220
REPOSITORIES: GEO
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Project description:Genome wide mapping of RNA polymearase III binding sites in Saccharomyces cerevisiae under normal growth and nutrient starved condition using ChIP-seq. Chromatin Immuno-precipitation (ChIP) was performed for FLAG tagged version of pol III subunit RPC128 after crosslinking the log-phase cells with formaldehyde. MOCK and IP DNA was sequenced and coverage of pol III was calculated at each base of the genome. RPC128-FLAG ChIP-seq single end seqquencing on Illumina GAII. 2 replicates of IP samples and 1 MOCK sample. Done in under normal growth and nutrient deprivation (4 hours).
2013-06-21 | E-GEOD-39566 | biostudies-arrayexpress
Project description:Purpose: Characterization of the target genes of two versions of the Polymerase III (pol III), one containing POLR3G subunit or POLR3GL subunit, in different tissues. Methods: We performed ChIP_seq of POLR3D and the two variants subunits, POLR3G and POLR3GL in Human IMR90 cells, Mouse liver and Mouse Hepatocarcinoma Hepa 1-6 cells in order to locate and analyse total pol III, POLR3G and POLR3GL-containing pol III in various tissues. Results: We show that POLR3G- as well as POLR3GL-containing pol III are present in cultured cell lines and in normal mouse liver, although the relative amounts of the two forms vary, with the POLR3G-containing pol III relatively more abundant in dividing cells. Both forms of pol III occupy the same target genes, in very constant proportions within one cell line, suggesting that the two forms of pol III have similar function with regard to specificity for target genes. Conclusions: POLR3G and POLR3GL occupy the same target genes, but they are differentially expressed under different conditions and in different cell types. Pol III target genes in human IMR cells, Mouse liver, and Mouse Hepatocarcinoma cells were identified by chromatin Immuno-precipitation followed by deep sequencing using Illumina HiSeq. For Human IMR90 cells, we used antibodies against POLR3D, POLR3G, POLR3GL and BDP1. For Mouse data, we targeted POLR3D, POLR3G and POLR3GL and did ChIP_seq replicates. We also sequenced the corresponding inputs (crosslinked DNA of IMR90 cells, Mouse liver, and Mouse Hepatocarcinoma cells). This represents a total of 20 samples.
2013-10-01 | E-GEOD-47849 | biostudies-arrayexpress
Project description:In higher eukaryotes, an important mechanism to tune translation in different tissues and conditions is mTORC1-dependent regulation of tRNAs transcription by RNA polymerase III (Pol III), as the relative amount of tRNAs tightly coordinates with the translational needs of the cell. mTORC1 contributes to regulate protein synthesis through its direct substrate MAF1, which functions as a negative regulator of Pol III transcription in response to stimuli such as serum starvation or rapamycin treatment. Here, we applied ChIP-seq to examine the Pol III occupancy profile in human fibroblasts and report evidence of a genome wide, MAF1-dependent coordinated response to favorable or stress growth conditions. Strikingly, while a set of genes is extremely responsive in terms of Pol III binding, other genes are mostly unperturbed, yet associated with transcriptionally engaged polymerases as revealed by nascent EU-labeled RNA-seq (neuRNA-seq). As shown by DamIP-seq, the responsiveness of a subset of genes is tightly connected to the rapid and transient interaction of MAF1 with DNA-bound Pol III. We performed duplicate ChIP-seq experiments for the Rpc4 (POLR3D) subunit of RNA polymerase III in IMR90hTert cells grown in the presence of fetal bovine serum (FBS), serum starved (SS), serum starved and treated with insulin (SS+I), serum starved and treated with insulin and rapamycin (SS+R+I). Additional ChIP-seq profiles were generated in cells treated with MAF1 siRNAs and serum starved. MAF1 binding was addressed by DamIP-seq, using two replicates per clone of IMR90hTert cells expressing HA-tagged MAF1-DamK9A (2 different clones) or EGFP-DamK9A (2 different clones). To monitor dynamic transcription profiles we did neusRNA-seq in IMR90hTert cells EU-labeled or mock (DMSO)-labeled. For both DamIP-seq and neusRNA-seq, cells were either unperturbed or serum starved.
2016-02-15 | E-GEOD-73928 | biostudies-arrayexpress
Project description:In higher eukaryotes, an important mechanism to tune translation in different tissues and conditions is mTORC1-dependent regulation of tRNAs transcription by RNA polymerase III (Pol III), as the relative amount of tRNAs tightly coordinates with the translational needs of the cell. mTORC1 contributes to regulate protein synthesis through its direct substrate MAF1, which functions as a negative regulator of Pol III transcription in response to stimuli such as serum starvation or rapamycin treatment. Here, we applied ChIP-seq to examine the Pol III occupancy profile in human fibroblasts and report evidence of a genome wide, MAF1-dependent coordinated response to favorable or stress growth conditions. Strikingly, while a set of genes is extremely responsive in terms of Pol III binding, other genes are mostly unperturbed, yet associated with transcriptionally engaged polymerases as revealed by nascent EU-labeled RNA-seq (neuRNA-seq). As shown by DamIP-seq, the responsiveness of a subset of genes is tightly connected to the rapid and transient interaction of MAF1 with DNA-bound Pol III. We performed duplicate ChIP-seq experiments for the Rpc4 (POLR3D) subunit of RNA polymerase III in IMR90hTert cells grown in the presence of fetal bovine serum (FBS), serum starved (SS), serum starved and treated with insulin (SS+I), serum starved and treated with insulin and rapamycin (SS+R+I). Additional ChIP-seq profiles were generated in cells treated with MAF1 siRNAs and serum starved. MAF1 binding was addressed by DamIP-seq, using two replicates per clone of IMR90hTert cells expressing HA-tagged MAF1-DamK9A (2 different clones) or EGFP-DamK9A (2 different clones). To monitor dynamic transcription profiles we did neusRNA-seq in IMR90hTert cells EU-labeled or mock (DMSO)-labeled. For both DamIP-seq and neusRNA-seq, cells were either unperturbed or serum starved.
2016-02-15 | E-GEOD-73926 | biostudies-arrayexpress
Project description:In higher eukaryotes, an important mechanism to tune translation in different tissues and conditions is mTORC1-dependent regulation of tRNAs transcription by RNA polymerase III (Pol III), as the relative amount of tRNAs tightly coordinates with the translational needs of the cell. mTORC1 contributes to regulate protein synthesis through its direct substrate MAF1, which functions as a negative regulator of Pol III transcription in response to stimuli such as serum starvation or rapamycin treatment. Here, we applied ChIP-seq to examine the Pol III occupancy profile in human fibroblasts and report evidence of a genome wide, MAF1-dependent coordinated response to favorable or stress growth conditions. Strikingly, while a set of genes is extremely responsive in terms of Pol III binding, other genes are mostly unperturbed, yet associated with transcriptionally engaged polymerases as revealed by nascent EU-labeled RNA-seq (neuRNA-seq). As shown by DamIP-seq, the responsiveness of a subset of genes is tightly connected to the rapid and transient interaction of MAF1 with DNA-bound Pol III. We performed duplicate ChIP-seq experiments for the Rpc4 (POLR3D) subunit of RNA polymerase III in IMR90hTert cells grown in the presence of fetal bovine serum (FBS), serum starved (SS), serum starved and treated with insulin (SS+I), serum starved and treated with insulin and rapamycin (SS+R+I). Additional ChIP-seq profiles were generated in cells treated with MAF1 siRNAs and serum starved. MAF1 binding was addressed by DamIP-seq, using two replicates per clone of IMR90hTert cells expressing HA-tagged MAF1-DamK9A (2 different clones) or EGFP-DamK9A (2 different clones). To monitor dynamic transcription profiles we did neusRNA-seq in IMR90hTert cells EU-labeled or mock (DMSO)-labeled. For both DamIP-seq and neusRNA-seq, cells were either unperturbed or serum starved.
2016-02-15 | E-GEOD-73929 | biostudies-arrayexpress
Project description:The genomic distribution of transcriptionally engaged Pol II in control and heat shocked cells was determined by combining formaldehyde crosslinking and permanganate oxidation of transcription bubbles Cells treated in succession with formaldehyde and then permanganate, were subjected to chromatin precipitation with Rpb3 antibody, and then the pattern of permanganate modifications were mapped genome-wide
2013-07-02 | E-GEOD-46620 | biostudies-arrayexpress
Project description:Pol III ChIP-seq mouse development
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Project description:MAF1 represses Pol III-mediated transcription by interfering with TFIIIB and Pol III. Herein, we found that MAF1 knockdown induced CDKN1A transcription and chromatin looping concurrently with Pol III recruitment. Simultaneous knockdown of MAF1 with Pol III or BRF1 (subunit of TFIIIB) diminished the activation and looping effect, which indicates that recruiting Pol III was required for activation of Pol II-mediated transcription and chromatin looping. ChIP analysis after MAF1 knockdown indicated enhanced binding of Pol III and BRF1, as well as of CFP1, p300, and PCAF, which are factors that mediate active histone marks, along with the binding of TBP and POLR2E to the CDKN1A promoter. Simultaneous knockdown with Pol III abolished these regulatory events. Similar results were obtained for GDF15. Our results reveal a novel mechanism by which MAF1 and Pol III regulate the activity of a protein-coding gene transcribed by Pol II.
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