Genomics

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161

The conserved protein Seb1 drives transcription termination by binding RNA polymerase II and nascent RNA


ABSTRACT: Termination of Pol II transcription is an important step in the transcription cycle and is responsible for dislodgement of polymerase from DNA which leads to the release of a functional transcript. Recent studies have identified key players important for termination and showed a conserved domain that interacts with the phosphorylated C-terminus of Pol II (CTD-Interacting-Domain, CID) to constitute a common feature of these proteins. However, the mechanism by which transcription termination is achieved, is not understood. Using genome-wide methods, we demonstrate that the fission yeast CID protein Seb1 is essential for termination of protein-coding and non-coding genes through interacting with S2-phosphorylated Pol II and nascent RNA. Furthermore, we present the crystal structures of the Seb1 CTD- and RNA-binding modules. Unexpectedly, the latter reveals a novel intertwined two-domain arrangement of a canonical RRM and a second domain. These results provide important insights into the mechanism underlying eukaryotic transcription termination. Overall design: In this study, we studied binding of the S. pombe protein Seb1 directly to RNA by PAR-CLIP which was normalised to RNA expression levels (for which a RNA-Seq in WT cells was included). Point mutations were then introduced into Seb1 using a repressible system and RNA-Seq was performed for Seb1-WT and 5 different Seb1 point mutants that were all treated in the same way. Some samples were sequenced in duplicates and some (including the WT) in triplicates. Finally, binding was compared to another transcription factor, Pcf11, for which ChIP-Seq was performed. A control was done using an untagged strain. The binding profile was compared to phosphorylation levels of Ser2 on RNA-Pol II by doing ChIP-Seq for which an antibody against S2P was utilised. All ChIP-Seq experiments include an input sample.

INSTRUMENT(S): Ion Torrent PGM (Schizosaccharomyces pombe)

SUBMITTER: Sina Wittmann  

PROVIDER: GSE89914 | GEO | 2017-02-05

SECONDARY ACCESSION(S): PRJNA354266

REPOSITORIES: GEO

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The conserved protein Seb1 drives transcription termination by binding RNA polymerase II and nascent RNA.

Wittmann Sina S   Renner Max M   Watts Beth R BR   Adams Oliver O   Huseyin Miles M   Baejen Carlo C   El Omari Kamel K   Kilchert Cornelia C   Heo Dong-Hyuk DH   Kecman Tea T   Cramer Patrick P   Grimes Jonathan M JM   Vasiljeva Lidia L  

Nature communications 20170403


Termination of RNA polymerase II (Pol II) transcription is an important step in the transcription cycle, which involves the dislodgement of polymerase from DNA, leading to release of a functional transcript. Recent studies have identified the key players required for this process and showed that a common feature of these proteins is a conserved domain that interacts with the phosphorylated C-terminus of Pol II (CTD-interacting domain, CID). However, the mechanism by which transcription terminati  ...[more]

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