Dissecting the stochastic transcription initiation process in live Escherichia coli.
DNA Res
; 23(3): 203-14, 2016 Jun.
Article
in En
| MEDLINE
| ID: mdl-27026687
ABSTRACT
We investigate the hypothesis that, in Escherichia coli, while the concentration of RNA polymerases differs in different growth conditions, the fraction of RNA polymerases free for transcription remains approximately constant within a certain range of these conditions. After establishing this, we apply a standard model-fitting procedure to fully characterize the in vivo kinetics of the rate-limiting steps in transcription initiation of the Plac/ara-1 promoter from distributions of intervals between transcription events in cells with different RNA polymerase concentrations. We find that, under full induction, the closed complex lasts â¼788 s while subsequent steps last â¼193 s, on average. We then establish that the closed complex formation usually occurs multiple times prior to each successful initiation event. Furthermore, the promoter intermittently switches to an inactive state that, on average, lasts â¼87 s. This is shown to arise from the intermittent repression of the promoter by LacI. The methods employed here should be of use to resolve the rate-limiting steps governing the in vivo dynamics of initiation of prokaryotic promoters, similar to established steady-state assays to resolve the in vitro dynamics.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Escherichia coli
/
Transcription Initiation, Genetic
/
Models, Genetic
Language:
En
Journal:
DNA Res
Journal subject:
BIOLOGIA MOLECULAR
/
GENETICA
Year:
2016
Document type:
Article
Affiliation country:
Finland