Journal article
Mechanism of Transcriptional Activation by Pseudomonas aeruginosa ExsA
Journal of bacteriology, Vol.191(21), pp.6654-6664
11/2009
DOI: 10.1128/JB.00902-09
PMCID: PMC2795306
PMID: 19717612
Abstract
ExsA is a transcriptional activator of the
Pseudomonas aeruginosa
type III secretion system (T3SS). The T3SS consists of >40 genes organized within 10 transcriptional units, each of which is controlled by the transcriptional activator ExsA. ExsA-dependent promoters contain two adjacent ExsA binding sites that when occupied protect the −30 to −70 region from DNase I cleavage. The promoters also possess regions bearing strong resemblance to the consensus −10 and −35 regions of σ
70
-dependent promoters. The spacing distance between the putative −10 and −35 regions of ExsA-dependent promoters, however, is increased by 4 to 5 bp compared to that in typical σ
70
-dependent promoters. In the present study, we demonstrate that ExsA-dependent transcriptional activation requires a 21- or 22-bp spacer length between the −10 and −35 regions. Despite the atypical spacing in this region, in vitro transcription assays using σ
70
-saturated RNA polymerase holoenzyme (RNAP-σ
70
) confirm that ExsA-dependent promoters are indeed σ
70
dependent. Potassium permanganate footprinting experiments indicate that ExsA facilitates an early step in transcriptional initiation. Although RNAP-σ
70
binds to the promoters with low affinity in the absence of ExsA, the activator stimulates transcription by enhancing recruitment of RNAP-σ
70
to the P
exsC
and P
exsD
promoters. Abortive initiation assays confirm that ExsA enhances the equilibrium binding constant for RNAP while having only a modest effect on the isomerization rate constant.
Details
- Title: Subtitle
- Mechanism of Transcriptional Activation by Pseudomonas aeruginosa ExsA
- Creators
- Christopher A Vakulskas - Department of Microbiology, University of Iowa, Iowa City, IowaKeith M Brady - Department of Microbiology, University of Iowa, Iowa City, IowaTimothy L Yahr - Department of Microbiology, University of Iowa, Iowa City, Iowa
- Resource Type
- Journal article
- Publication Details
- Journal of bacteriology, Vol.191(21), pp.6654-6664
- DOI
- 10.1128/JB.00902-09
- PMID
- 19717612
- PMCID
- PMC2795306
- NLM abbreviation
- J Bacteriol
- ISSN
- 0021-9193
- eISSN
- 1098-5530
- Publisher
- American Society for Microbiology (ASM)
- Language
- English
- Date published
- 11/2009
- Academic Unit
- Microbiology and Immunology
- Record Identifier
- 9984001130102771
Metrics
19 Record Views