The Mot1 ATPase and Spt16 Histone Chaperone Co-Regulate Transcription Through Preinitiation Complex Assembly and Nucleosome Organization
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PROVIDER: PRJNA318368 | ENA |
REPOSITORIES: ENA
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Project description:Mot1 is a conserved and essential Swi2/Snf2 ATPase that can remove TATA-binding protein (TBP) from DNA using ATP hydrolysis, and in so doing exerts global effects on transcription. Spt16 is also essential and functions globally in transcriptional regulation as a component of the FACT histone chaperone complex. Here we demonstrate that Mot1 and Spt16 regulate a largely overlapping set of genes in Saccharomyces cerevisiae. As expected, Mot1 was found to control TBP levels at co-regulated promoters. In contrast, Spt16 did not affect TBP recruitment. Interestingly, Mot1 was required for Spt16 recruitment to co-activated promoters. In contrast, Spt16 levels in gene coding regions were unaffected by Mot1 as well as RNA polymerase II density. The co-localization of Mot1 and Spt16 at promoters and the broad overlap in the sets of genes they control is consistent with physical and genetic interactions between them. The data support a model in which these factors participate in a regulatory pathway in which Mot1 acts upstream of Spt16. Tiling arrays covering the entirety of the S.cerevisiae genome were used to identify the effects of Mot1 and Spt16 on RNA expression genome-wide. All samples were done in biological duplicates. The average signal from the spt16-197 samples was compared to the SPT16-WT to determine changes in expression. The average signal from the double mutant mot1-42 spt16-197 was compared to both SPT16-WT and MOT1-WT. The MOT1-WT data was previously published by our lab and is available at GEO accession GSM456548. Comparisons were made from our Spt16 dataset to the previously published MOT1-WT and mot1-42 data, and the entire study is available at GEO accession GSE18283. Differential RNA (spt16-197/SPT16): spt16-197_over_SPT16-WT.bar Differential RNA (mot1-42 spt16-197/SPT16): dbl_mut_over_SPT16-WT.bar Differential RNA (mot1-42 spt16-197/MOT1): dbl_mut_over_MOT1-WT.bar
2016-05-16 | E-GEOD-72117 | biostudies-arrayexpress
Project description:Mot1 is a conserved and essential Swi2/Snf2 ATPase that can remove TATA-binding protein (TBP) from DNA using ATP hydrolysis, and in so doing exerts global effects on transcription. Spt16 is also essential and functions globally in transcriptional regulation as a component of the FACT histone chaperone complex. Here we demonstrate that Mot1 and Spt16 regulate a largely overlapping set of genes in Saccharomyces cerevisiae. As expected, Mot1 was found to control TBP levels at co-regulated promoters. In contrast, Spt16 did not affect TBP recruitment. Interestingly, Mot1 was required for Spt16 recruitment to co-activated promoters. In contrast, Spt16 levels in gene coding regions were unaffected by Mot1 as well as RNA polymerase II density. The co-localization of Mot1 and Spt16 at promoters and the broad overlap in the sets of genes they control is consistent with physical and genetic interactions between them. The data support a model in which these factors participate in a regulatory pathway in which Mot1 acts upstream of Spt16.
2016-05-16 | GSE72117 | GEO
Project description:Mot1 is a conserved and essential Swi2/Snf2 ATPase that can remove TATA-binding protein (TBP) from DNA using ATP hydrolysis, and in so doing exerts global effects on transcription. Spt16 is also essential and functions globally in transcriptional regulation as a component of the FACT histone chaperone complex. Here we demonstrate that Mot1 and Spt16 regulate a largely overlapping set of genes in Saccharomyces cerevisiae. As expected, Mot1 was found to control TBP levels at co-regulated promoters. In contrast, Spt16 did not affect TBP recruitment. On a global scale, Spt16 was required for Mot1 promoter localization, and Mot1 also affected Spt16 localization to genes. Interestingly, we find that Mot1 has an unanticipated role in establishing or maintaining the occupancy and positioning of nucleosomes at the 5’ ends of genes. Spt16 has a broad role in regulating chromatin organization in gene bodies, including those nucleosomes affected by Mot1. These results suggest that the large-scale overlap in Mot1 and Spt16 function arises from a combination of both their unique and shared functions in transcription complex assembly and chromatin structure regulation.
2016-05-16 | GSE80234 | GEO
Project description:Mot1 is a conserved and essential Swi2/Snf2 ATPase that can remove TATA-binding protein (TBP) from DNA using ATP hydrolysis, and in so doing exerts global effects on transcription. Spt16 is also essential and functions globally in transcriptional regulation as a component of the FACT histone chaperone complex. Here we demonstrate that Mot1 and Spt16 regulate a largely overlapping set of genes in Saccharomyces cerevisiae. As expected, Mot1 was found to control TBP levels at co-regulated promoters. In contrast, Spt16 did not affect TBP recruitment. On a global scale, Spt16 was required for Mot1 promoter localization, and Mot1 also affected Spt16 localization to genes. Interestingly, we find that Mot1 has an unanticipated role in establishing or maintaining the occupancy and positioning of nucleosomes at the 5â ends of genes. Spt16 has a broad role in regulating chromatin organization in gene bodies, including those nucleosomes affected by Mot1. These results suggest that the large-scale overlap in Mot1 and Spt16 function arises from a combination of both their unique and shared functions in transcription complex assembly and chromatin structure regulation. ChIP was performed for Spt16-myc in WT cells and mot1-42 cells in duplicate with input DNA from WT as control. ChIP was performed for Mot1-myc in WT cells and spt16-197 cells in dublicate with input DNA from WT as control. Micrococcal nuclease digested chromatin from WT, mot1-42, spt16-197, and mot1-42 spt16-197 cells were immunoprecipitated with H3 antibody in duplicate. All samples were sequenced by Illumina MiSeq.
2016-05-16 | E-GEOD-80234 | biostudies-arrayexpress
Project description: Not available
| S-EPMC4946942 | biostudies-literature
Project description:FACT consists of two essential subunits Spt16 and Pob3 and is a histone chaperone. Depletion of Spt16 using a an spt16 mutant results in a global alteration of nucleosome positions as well as aberrant transcription. Here we show that the majority of nucleosomal changes at gene body upon Spt16 depletion are independent of gene activity, but correlates with cryptic gene transcription and are suppressed by inhibition of RNA Polymerase II activities. In addition, A small fraction of nucleosomal changes is resistant to Pol II inhibition, and Spt16 is enriched at this subgroup of nucleosomes. Moreover, nucleosomes surrounding the initiation sites of cryptic transcription in the spt16 mutant cells are more dynamic than other regions. These results support a model that Spt16 maintains nucleosome stability locally to inhibit the initiation from cryptic transcription, which that once initiated drives additional nucleosome loss upon Spt16 depletion.
2016-05-05 | GSE66215 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below.
2016-05-16 | GSE80235 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2016-05-16 | E-GEOD-80235 | biostudies-arrayexpress
Project description:TATA-binding protein (TBP) nucleates the assembly of the transcription preinitiation complex (PIC), and although TBP can bind promoters with high stability in vitro, recent results establish that virtually the entire TBP population is highly dynamic in yeast nuclei in vivo. This dynamic behavior is surprising in light of models which posit that a stable TBP-containing scaffold facilitates transcription reinitiation at active promoters. The dynamic behavior of TBP is a consequence of the enzymatic activity of the essential Snf2/Swi2 ATPase Mot1, suggesting that ensuring a highly mobile TBP population is critical for transcriptional regulation on a global scale. Here high-resolution tiling arrays were used to define how perturbed TBP dynamics impact the precision of RNA synthesis in Saccharomyces cerevisiae. We find that Mot1 plays a broad role in establishing the precision and efficiency of RNA synthesis: in mot1-42 cells, RNA length changes were observed for 713 genes, about twice the number observed in set2∆ cells, which display a previously reported propensity for spurious initiation within open reading frames. Loss of Mot1 led to both aberrant transcription initiation and termination, with prematurely terminated transcripts representing the largest class of events. Genetic and genomic analyses support the conclusion that these effects on RNA length are mechanistically tied to dynamic TBP occupancies at certain types of promoters. These results suggest a new model whereby dynamic disassembly of the PIC can influence productive RNA synthesis.
2010-09-08 | GSE18283 | GEO
Project description:The heterodimeric histone chaperone FACT consisting of SSRP1 and SPT16 contributes to dynamic nucleosome rearrangements during various DNA-dependent processes including transcription. In search of post-translational modifications that may regulate the activity of FACT, SSRP1 and SPT16 were isolated from Arabidopsis cells and analysed by mass spectrometry. Four acetylated lysine residues could be mapped within the basic C-terminal region of SSRP1, while three phosphorylated serine/threonine residues were identified in the acidic C-terminal region of SPT16. Mutational analysis of the SSRP1 acetylation sites revealed only mild effects. However, phosphorylation of SPT16 that is catalysed by protein kinase CK2, modulates histone interactions. A non-phosphorylatable version of SPT16 displayed reduced histone binding and (unlike the phosphorylatable wild-type and phosphomimic versions) proved inactive in complementing the growth and developmental phenotypes of spt16 mutant plants. In plants expressing the non-phosphorylatable SPT16 version we detected at a subset of genes enrichment of histone H3 directly upstream of RNA polymerase II transcriptional start sites (TSSs) in a region that usually is nucleosome-depleted. This is associated with altered transcript levels, suggesting that some genes require phosphorylation of the SPT16 acidic region for establishing the correct nucleosome occupancy at the TSS as a prerequisite for proper transcription.
2022-06-09 | PXD028534 | Pride