Altering transcription factor binding reveals comprehensive transcriptional kinetics of a basic gene
| Autor: | Achim P. Popp, J. Christof M. Gebhardt, Johannes Hettich |
|---|---|
| Jazyk: | angličtina |
| Předmět: |
Transcriptional Activation
Transcription Genetic AcademicSubjects/SCI00010 RNA polymerase II Biology Cell Line 03 medical and health sciences 0302 clinical medicine Transcription (biology) Genes Reporter Genetics Transcription factor Gene Transcription Activator-Like Effectors 030304 developmental biology Regulation of gene expression 0303 health sciences Messenger RNA Reporter gene Gene regulation Chromatin and Epigenetics DNA Cell biology Chromatin Kinetics biology.protein 030217 neurology & neurosurgery Transcription Factors |
| Zdroj: | Nucleic Acids Research |
| ISSN: | 1362-4962 0305-1048 |
| DOI: | 10.1093/nar/gkab443 |
| Popis: | Transcription is a vital process activated by transcription factor (TF) binding. The active gene releases a burst of transcripts before turning inactive again. While the basic course of transcription is well understood, it is unclear how binding of a TF affects the frequency, duration and size of a transcriptional burst. We systematically varied the residence time and concentration of a synthetic TF and characterized the transcription of a synthetic reporter gene by combining single molecule imaging, single molecule RNA-FISH, live transcript visualisation and analysis with a novel algorithm, Burst Inference from mRNA Distributions (BIRD). For this well-defined system, we found that TF binding solely affected burst frequency and variations in TF residence time had a stronger influence than variations in concentration. This enabled us to device a model of gene transcription, in which TF binding triggers multiple successive steps before the gene transits to the active state and actual mRNA synthesis is decoupled from TF presence. We quantified all transition times of the TF and the gene, including the TF search time and the delay between TF binding and the onset of transcription. Our quantitative measurements and analysis revealed detailed kinetic insight, which may serve as basis for a bottom-up understanding of gene regulation. |
| Databáze: | OpenAIRE |
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