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| Status |
Public on Dec 20, 2021 |
| Title |
The interaction between the Spt6-tSH2 domain and Rpb1 affects multiple functions of RNA Polymerase II [MNase-seq] |
| Organism |
Saccharomyces cerevisiae |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
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.
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| Overall design |
6 samples (3 replicates each of 2 yeast strains) were grown to logarithmic phase (final OD ~0.8) in rich medium at 30° C. Cells were crosslinked with 1% formaldehyde for 20 minutes, collected by centrifugation, spheroplasted, digested with Micrococcal nuclease, then DNA was extracted.
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| Contributor(s) |
Connell Z, McCullough LL, Parnell TJ, Formosa T |
| Citation(s) |
34967414 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 GM064649 |
Biochemical, Genetic, and Genomic Analysis of Nucleosome Reorganization by FACT |
UNIVERSITY OF UTAH |
Timothy G Formosa |
| R01 GM116560 |
Structure, mechanism, and function of the histone chaperones Spt6 and FACT |
UNIVERSITY OF UTAH |
Timothy G Formosa |
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| Submission date |
Nov 29, 2021 |
| Last update date |
Feb 02, 2022 |
| Contact name |
Tim Formosa |
| E-mail(s) |
[email protected]
|
| Phone |
8015815435
|
| Organization name |
University of Utah School of Medicine
|
| Department |
Biochemistry
|
| Street address |
15 N Medical Dr East RM 4100
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| City |
Salt Lake City |
| State/province |
UT |
| ZIP/Postal code |
84112-5650 |
| Country |
USA |
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| Platforms (1) |
| GPL27812 |
Illumina NovaSeq 6000 (Saccharomyces cerevisiae) |
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| Samples (6)
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| This SubSeries is part of SuperSeries: |
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| Relations |
| BioProject |
PRJNA784614 |
| SRA |
SRP348497 |
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