Spt6 association with RNAPII directs mRNA turnover during transcription
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA438236 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6833140_1.fastq.gz | Fastqsanger.gz | |||
| SRR6833140_2.fastq.gz | Fastqsanger.gz | |||
| SRR6833141_1.fastq.gz | Fastqsanger.gz | |||
| SRR6833141_2.fastq.gz | Fastqsanger.gz | |||
| SRR6833132.fastq.gz | Fastqsanger.gz |
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Similar Datasets
Project description:Spt6 is an essential histone chaperone that mediates nucleosome reassembly during gene transcription. Spt6 interacts with elongating RNA polymerase II (RNAPII) via a tandem Src2 homology (tSH2) domain, but it is not known whether this particular interaction is required for the nucleosome reassembly activity of Spt6. Here, we show that Spt6 recruitment to genes and its nucleosome reassembly functions are largely independent of association with RNAPII. Instead, the Spt6-RNAPII association is required for post-transcriptional mRNA turnover. Mechanistically, association of Spt6 with RNAPII couples the Ccr4-Not complex to the transcribed regions of genes, which we show regulates the timely deadenylation and degradation of a broad range of mRNAs including those required for cell cycle progression. Thus, our findings reveal an unexpected control mechanism for mRNA turnover facilitated by a histone chaperone during transcription.
2018-04-18 | GSE111815 | GEO
Project description:Spt6-Spn1 is an essential histone chaperone complex that associates with RNA Polymerase II (RNAPII) and reassembles nucleosomes during gene transcription. While Spt6-Spn1 interaction is important to its histone deposition and transcription functions, a precise understanding of how Spt6-Spn1 association contributes to its functions is still limited. Here, using temperature sensitive alleles of spt6 and spn1 that disrupt their interaction, we show that Spt6-Spn1 association is broadly important for its interaction with the elongating RNAPII complex and nucleosomes. Using micrococcal nuclease (MNase)-based chromatin occupancy profiling, we further find that Spt6-Spn1 interaction is required for maintaining the preferred nucleosome positioning generated from replication-dependent histone deposition at actively transcribed genes. These analyses also reveal an overall shift of nucleosomes towards the 5’ end of genes that correlates with decreased RNAPII levels. While loss of Spt6 and Spn1 function is known to promote cryptic transcription, the nucleosome defects found in spt6 and spn1 mutants that interfere with their interaction were observed broadly across transcribed genes, further highlighting the role of DNA sequence as a mediator of cryptic transcription when nucleosomes positioning is altered. Taken together, these findings reveal Spt6-Spn1 interaction is key to its association with elongating RNAPII and for its ability to precisely organize nucleosomes across transcription units.
2025-04-09 | GSE284959 | GEO
Project description:The histone chaperone Spt6 is involved in promoting elongation of RNA polymerase II (RNAPII), maintaining chromatin structure, regulating co-transcriptional histone modifications, and controlling mRNA processing. These diverse functions of Spt6 are partly mediated through its interactions with RNAPII and other factors in the transcription elongation complex. In this study, we used mass spectrometry to characterize the differences in RNAPII interacting factors between wild-type cells and those depleted for Spt6, leading to the identification of proteins that depend on Spt6 for their interaction with RNAPII. In all, eight samples were processed - four genotypes (1. SPT6, RPB3-untagged; 2. SPT6, RPB3-tagged; 3. spt6-1004, RPB3-untagged; 4. spt6-1004; RPB3-tagged) in biological duplicates.
2021-12-05 | PXD027708 | Pride
Project description:The histone chaperone Spt6 is involved in promoting elongation of RNA polymerase II (RNAPII), maintaining chromatin structure, regulating co-transcriptional histone modifications, and controlling mRNA processing. These diverse functions of Spt6 are partly mediated through its interactions with RNAPII and other factors in the transcription elongation complex. In this study, we used mass spectrometry to characterize the differences in RNAPII interacting factors between wild-type cells and those depleted for Spt6, leading to the identification of proteins that depend on Spt6 for their interaction with RNAPII. The altered association of some of these factors could be attributed to changes in steady-state protein levels. However, Abd1, the mRNA cap methyltransferase, had decreased association with RNAPII after Spt6 depletion despite unchanged Abd1 protein levels, showing a requirement for Spt6 in mediating the Abd1-RNAPII interaction. Genome-wide studies showed that Spt6 is required for maintaining the level of Abd1 over transcribed regions, as well as the level of Spt5, another protein known to recruit Abd1 to chromatin. Abd1 levels were particularly decreased at the 5’ ends of genes after Spt6 depletion, suggesting a greater need for Spt6 in Abd1 recruitment over these regions. Together, our results show that Spt6 is important in regulating the composition of the transcription elongation complex and reveal a previously unknown function for Spt6 in the recruitment of Abd1.
2021-09-10 | GSE171953 | GEO
Project description:Spt6 is an essential histone chaperone that mediates nucleosome reassembly during gene transcription. Spt6 also associates with RNA polymerase II (RNAPII) via a tandem Src2 homology domain. However, the significance of Spt6-RNAPII interaction is not well understood. Here, we show that Spt6 recruitment to genes and the nucleosome reassembly functions of Spt6 can still occur in the absence of its association with RNAPII. Surprisingly, we found that Spt6-RNAPII association is required for efficient recruitment of the Ccr4-Not de-adenylation complex to transcribed genes for essential degradation of a range of mRNAs, including mRNAs required for cell-cycle progression. These findings reveal an unexpected control mechanism for mRNA turnover during transcription facilitated by a histone chaperone.
| S-EPMC6323640 | biostudies-literature
Project description:Spt6 is a multifunctional histone chaperone involved in the maintenance of chromatin structure during elongation by RNA polymerase II (Pol II). Spt6 has a tandem SH2 (tSH2) domain within its C-terminus that recognizes Pol II CTD peptides phosphorylated on Ser2, Ser5 or Try1 in vitro. Deleting the tSH2 domain, however, only has a partial effect on Spt6 occupancy in vivo, suggesting that more complex mechanisms are involved in the Spt6 recruitment. Our results show that the Ser2 kinases Bur1 and Ctk1, but not the Ser5 kinase Kin28, cooperate in recruiting Spt6, genome-wide. Interestingly, the Ser2 kinases promote the association of Spt6 in early transcribed regions and not toward the 3' end of genes, where phosphorylated Ser2 reaches its maximum level. Additionally, our results uncover an unexpected role for histone deacetylases (Rpd3 and Hos2) in promoting Spt6 interaction with elongating Pol II. Finally, our data suggest that phosphorylation of the Pol II CTD on Tyr1 promotes the association of Spt6 with the 3' end of transcribed genes, independently of Ser2 phosphorylation. Collectively, our results show that a complex network of interactions, involving the Spt6 tSH2 domain, CTD phosphorylation and histone deacetylases, coordinate the recruitment of Spt6 to transcribed genes in vivo. We examined the genome-wide distribution (using ChIP-chip) of Spt6. Spt6 occupancy was also assayed in mutants for CTD Serine 2 and Serine 5 kinases and in mutants for histone deacetylases. ChIPs were performed with a Myc-tagged version of Spt6. Most ChIPs (in Cy5) were hybridyzed against a control ChIP sample from an isogenic non-tagged strain (in Cy3). In the ChIP experiments with the spt6-202del mutant, non immunoprecipitated DNA (input) was used as the control. In addition to Spt6 ChIPs, the project includes RNAPII (Rpb3) ChIP-chip datasets, where an anti-Rpb3 antibody was used to ChIP RNAPII and non immunoprecipitated DNA (input) was used as the control. All ChIP-chip experiments were done in duplicates. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).
2014-09-25 | E-GEOD-61713 | biostudies-arrayexpress
Project description:Spt6 is a highly conserved factor that carries out important functions in transcription and chromatin structure. To gain new insights into Spt6, we measured nucleosome occupancy along Saccharomyces cerevisiae chromosome III in an spt6 mutant and found that the level of nucleosomes is greatly reduced accross some but not all coding regions. In addition, genome-wide location analyses of RNA polymerase II showed that the nucleosome loss in the spt6 mutant occurs over highly-transcribed genes. Unexpectedly, the effects of the spt6 mutation on nucleosome levels did not correlate with its effects on mRNA levels, suggesting that Spt6 plays distinct roles in controlling chromatin structure across coding regions and in transcriptional regulation. We studied one case of transcriptional regulation by Stp6, at the CHA1 gene, and showed that regulation likely occurs by Spt6 controlling the position of the +1 nucleosome. These results, along with previous studies, suggest that Spt6 regulates transcription by controlling chromatin structure over regulatory regions, and its effects on nucleosome levels over coding regions likely serve and independent function. In order to examine the genome-wide localization of RNAPII and Spt6 in Saccharomyces cerevisiae, RNAPII and Spt6 along with associated DNA sequences were immunoprecipitated using anti-8WG16 and anti-HA antibodies, respectively. The RNAPII and Spt6 chromatin immunoprecipitation was performed in duplicate from WT cells as described below. The extracted DNA was hybridized to a DNA microarray containing an average of 4 probes per kilobase across the whole yeast genome. The combined datasets are available in the supplemental files of the related publication.
2010-12-01 | E-GEOD-21787 | biostudies-arrayexpress
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:The histone chaperone and transcription elongation factor SPT6, integral to RNA Polymerase II (RNAPII) activity, also plays a crucial role in regulating transcription termination. In an attempt to identify the pathways employed by SPT6 in this regulation, we found that while SPT6 and its closest partner IWS1 interact and co-localize with RNAPII, their functions diverge significantly at gene termination sites. Our data revealed that SPT6 depletion, unlike IWS1, results in extensive readthrough transcription, indicating that SPT6 independently regulates transcription termination. Further analysis identified that the cleavage and polyadenylation factor PCF11 and the phosphatase regulatory protein PNUTS collaborate with SPT6 in this process. These findings suggest that SPT6 may facilitate transcription termination by recruiting PNUTS and PCF11 to RNAPII. Additionally, SPT6 and PNUTS jointly restrict promoter upstream transcripts (PROMPTs), whereas PCF11 presence is essential for their activation in the absence of SPT6. This study elucidates the distinct and overlapping functions of SPT6, PCF11, and PNUTS in transcription termination, highlighting the pivotal role of SPT6 in ensuring proper transcription termination at the 5’ and 3’ ends of genes.
2025-03-14 | GSE272847 | GEO
Project description:The histone chaperone and transcription elongation factor SPT6, integral to RNA Polymerase II (RNAPII) activity, also plays a crucial role in regulating transcription termination. In an attempt to identify the pathways employed by SPT6 in this regulation, we found that while SPT6 and its closest partner IWS1 interact and co-localize with RNAPII, their functions diverge significantly at gene termination sites. Our data revealed that SPT6 depletion, unlike IWS1, results in extensive readthrough transcription, indicating that SPT6 independently regulates transcription termination. Further analysis identified that the cleavage and polyadenylation factor PCF11 and the phosphatase regulatory protein PNUTS collaborate with SPT6 in this process. These findings suggest that SPT6 may facilitate transcription termination by recruiting PNUTS and PCF11 to RNAPII. Additionally, SPT6 and PNUTS jointly restrict promoter upstream transcripts (PROMPTs), whereas PCF11 presence is essential for their activation in the absence of SPT6. This study elucidates the distinct and overlapping functions of SPT6, PCF11, and PNUTS in transcription termination, highlighting the pivotal role of SPT6 in ensuring proper transcription termination at the 5’ and 3’ ends of genes.
2025-03-14 | GSE272848 | GEO