Requirements for RNA polymerase II preinitiation complex formation in vivo
- Harvard Medical School, United States
- University of Macau, Macau
Abstract
Transcription by RNA polymerase II requires assembly of a preinitiation complex (PIC) composed of general transcription factors (GTFs) bound at the promoter. In vitro, some GTFs are essential for transcription, whereas others are not required under certain conditions. PICs are stable in the absence of nucleotide triphosphates, and subsets of GTFs can form partial PICs. By depleting individual GTFs in yeast cells, we show that all GTFs are essential for TBP binding and transcription, suggesting that partial PICs do not exist at appreciable levels in vivo. Depletion of FACT, a histone chaperone that travels with elongating Pol II, strongly reduces PIC formation and transcription. In contrast, TBP-associated factors (TAFs) contribute to transcription of most genes, but TAF-independent transcription occurs at substantial levels, preferentially at promoters containing TATA elements. PICs are absent in cells deprived of uracil, and presumably UTP, suggesting that transcriptionally inactive PICs are removed from promoters in vivo.
Data availability
Sequencing data has been deposited in GEO under the accession number GSE122734
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Requirements for RNA polymerase II preinitiation complex formation in vivoNCBI Gene Expression Omnibus, GSE122734.
Article and author information
Author details
Funding
National Institutes of Health (GM 30186)
- Natalia Petrenko
- Yi Jin
- Koon Ho Wong
- Kevin Struhl
Universidade de Macau (MYRG2015-00186 FHS)
- Liguo Dong
- Koon Ho Wong
Croucher Foundation
- Koon Ho Wong
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Petrenko et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Requirements for RNA polymerase II preinitiation complex formation in vivo
eLife 8:e43654.
https://doi.org/10.7554/eLife.43654
Further reading
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Specialized magnetic beads that bind target proteins to a cryogenic electron microscopy grid make it possible to study the structure of protein complexes from dilute samples.
