Genome-wide maps of Spt6 occupancy in yeast strains with mutations in Spt6 and Rpb1
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA385007 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:We determined that the tandem SH2 domain of S. cerevisae Spt6 binds the linker region of the RNA polymerase II subunit Rpb1, rather than the expected sites in its heptad repeat domain. The 4 nM binding affinity requires phosphorylation at Rpb1 S1493 and either T1471 or Y1473. Crystal structures showed that pT1471 binds the canonical SH2 pY site while pS1493 binds an unanticipated pocket 70 Å distant. Remarkably, the pT1471 phosphate occupies the phosphate-binding site of a canonical pY complex, while Y1473 occupies the position of a canonical pY side chain, with the combination of pT and Y mimicking a pY moiety. Biochemical data and modeling indicate that pY1473 can form an equivalent interaction, and we find that pT1471/pS1493 and pY1473/pS1493 combinations occur in vivo. ChIP-seq and genetic analyses demonstrate the importance of these interactions for recruitment of Spt6 to sites of transcription and for the maintenance of repressive chromatin.
2017-10-24 | GSE98405 | GEO
Project description:The conserved transcription elongation factor Spt6 makes several contacts with the RNA Polymerase II (RNAPII) complex, including a high-affinity interaction between the Spt6 tandem SH2 domain (Spt6-tSH2) and phosphorylated residues of the Rpb1 subunit in a region linking the catalytic core and the heptad repeats at the C-terminal domain (CTD). This interaction contributes to generic localization of Spt6, but we show here that it also has gene-specific roles. Disrupting the interface affected transcription start site selection at a subset of genes whose expression is regulated by this choice, and this was accompanied by changes in the distinct pattern of Spt6 occupancy at these sites. Splicing efficiency was also impacted, as was apparent progression through introns that encode snoRNAs. Expression of genes that are particularly sensitive to efficient restoration of chromatin after transcription were affected, and a distinct role in maintaining +1 nucleosomes was identified, especially at ribosomal protein genes. The Spt6-tSH2:Rpb1 interface therefore has both genome-wide functions and local roles at subsets of genes where dynamic decisions regarding initiation, transcript processing, or termination are made. We propose that the Spt6-tSH2:Rpb1 interaction participates in coordinating appropriate responses to the varying local conditions encountered RNAPII at each gene, perhaps by modulating the elongation rate to allow reconfiguration of accessory factor activity or availability.
2021-12-20 | GSE184954 | GEO
Project description:The conserved transcription elongation factor Spt6 makes several contacts with the RNA Polymerase II (RNAPII) complex, including a high-affinity interaction between the Spt6 tandem SH2 domain (Spt6-tSH2) and phosphorylated residues of the Rpb1 subunit in the linker between the catalytic core and the C-terminal domain (CTD) heptad repeats. This interaction contributes to generic localization of Spt6, but we show here that it also has gene-specific roles. Disrupting the interface affected transcription start site selection at a subset of genes whose expression is regulated by this choice, and this was accompanied by changes in a distinct pattern of Spt6 accumulation at these sites. Splicing efficiency was also diminished, as was apparent progression through introns that encode snoRNAs. Chromatin-mediated repression was impaired, and a distinct role in maintaining +1 nucleosomes was identified, especially at ribosomal protein genes. The Spt6-tSH2:Rpb1 interface therefore has both genome-wide functions and local roles at subsets of genes where dynamic decisions regarding initiation, transcript processing, or termination are made. We propose that the interaction modulates the availability or activity of the core elongation and histone chaperone functions of Spt6, contributing to coordination between RNAPII and its accessory factors as varying local conditions call for dynamic responses.
2021-12-20 | GSE189831 | GEO
Project description:The conserved transcription elongation factor Spt6 makes several contacts with the RNA Polymerase II (RNAPII) complex, including a high-affinity interaction between the Spt6 tandem SH2 domain (Spt6-tSH2) and phosphorylated residues in Rpb1 in a region between the catalytic core and the heptad repeats of its C-terminal domain (CTD). This interaction contributes to the global occupancy of Spt6 within transcription units, suggesting that it has a general role in tethering Spt6 to the elongation complex. However, we show here that disrupting this binding caused increases in some transcripts, revealing specific functional roles in regulating the expression of subsets of genes. These included loci whose regulation involves differential transcription start site selection, early termination of transcription, or efficient restoration of chromatin integrity after transcription. Loss of this interaction also caused a defect in splicing, and apparent pausing of RNAPII progression in regions requiring more complex processing of excised introns. The results support a global role for the Spt6-tSH2:Rpb1 interaction as one of several means of stabilizing the association of Spt6 with RNAPII, but they also reveal local functions at specific sites, especially those where dynamic decisions regarding initiation or termination are made, or where changes in the configuration of associated factors occur. We therefore propose that the Spt6-tSH2:Rpb1 interaction can provide a conduit for communication between RNAPII and the elongation factor function of Spt6, or with other factors associated with the Rpb1 CTD, supporting appropriate elongation through challenging templates and efficient co-transcriptional processing.
2021-12-20 | GSE174427 | GEO
Project description:In fission yeast, the nuclear-localized Lsk1p-Lsc1p-Lsg1p cyclin dependent kinase complex is required for the reliable execution of cytokinesis and is also required for Ser-2 phosphorylation RNA pol II carboxy terminal domain. To address whether alterations in CTD phosphorylation might selectively alter expression of cytokinesis genes, expression profiling of site-directed CTD mutants was performed. Strains bearing the rpb1-12XCTD and rpb1-12XS2ACTD mutations were grown to mid-log phase in YES media and treated with 0.5uM LatA (or the solvent control, DMSO) for three hours at 30C. Three biological replicates were performed.
2011-08-31 | E-GEOD-31369 | biostudies-arrayexpress
Project description:To investigate the chromatin transcription cycle, we determined genome-wide occupancy profiles for RNA polymerase (Pol) II, its phosphorylated forms, and transcription factors in growing yeast. ChIP-chip was performed to identify the genomic binding locations for Rpb3, TFIIB, Tfg1, Kin28, Cet1, Spt4, Spt5, Spt6, Elf1, Spn1, Bur1, Ctk1, Paf1, Spt16, Pcf11, and Rpb1 phosphorylated at serine 2, 5, and 7 residues of the CTD, respectively.
2010-08-23 | E-TABM-1033 | biostudies-arrayexpress
Project description:Spt6 is a highly conserved histone chaperone that interacts directly with both RNA polymerase II and histones to regulate gene expression. To gain a comprehensive understanding of the requirements for this critical factor, we have performed genome-wide analyses of transcription, chromatin structure, and histone modifications in an S. pombe spt6 mutant. Our results demonstrate several dramatic changes to transcription and chromatin structure in the spt6 mutant, including an elevation of antisense transcripts at over 70 percent of all genes and general loss of the +1 nucleosome. Furthermore, Spt6 is required for the trimethylation of histone H3 on lysines 4 and 36, marks associated with active transcription. Taken together, our results indicate that Spt6 is critical for the accuracy of transcription and the integrity of chromatin, likely via its direct interactions with RNA polymerase II and histones. ChIP-seq experiments were performed on wild type and spt6-1 strains on the following proteins: RNA polymerase II (Rpb1), Paf1 Complex (Ctr9), COMPASS (Swd1), Set2, Spt6, histones H2B and H3, histone modifications H3K4me3 and H3K36me3. Experiments were performed in replicates and matching inputs were also sequenced.
2013-10-09 | E-GEOD-49574 | biostudies-arrayexpress
Project description:Spt6 is a conserved factor, critically required for several transcription and chromatin related processes. We now show that Spt6 and its binding partner, Iws1, are required for heterochromatic silencing in Schizosaccharomyces pombe. Our studies demonstrate that Spt6 is required for silencing of all heterochromatic loci and that an spt6 mutant has an unusual combination of heterochromatic phenotypes compared to previously studied silencing mutants. Unexpectedly, we find normal nucleosome positioning over heterochromatin and normal levels of histone H3K9 dimethylation. However, we also find greatly reduced levels of H3K9 trimethylation, elevated levels of H3K14 acetylation, and reduced recruitment of several silencing factors. Our evidence suggests that Spt6 plays a role at both the transcriptional and post-transcriptional levels; in an spt6 mutant, RNA polymerase II (RNAPII) occupancy at the pericentric regions is only modestly increased, while production of small interfering RNAs (siRNAs) is lost. Taken together, our results suggest that Spt6 is required for multiple steps in heterochromatic silencing by controlling chromatin, transcriptional, and post-transcriptional processes.
2011-10-20 | E-MTAB-738 | biostudies-arrayexpress
Project description:whole genome analysis of RNA pol II and histone H3 in WT and Spt6-depleted cells using a tetracycline regulated ts degron mutant, spt6-td.
2013-08-16 | GSE49926 | GEO