Abstract
Two-component signaling systems are a major strategy employed by bacteria, and to some extent, yeast and plants, to respond to environmental stress. The EnvZ/OmpR system in E. coli responds to osmotic and acid stress and is responsible for regulating the protein composition of the outer membrane. EnvZ is a histidine kinase located in the inner membrane. Upon activation, it is autophosphorylated by ATP and subsequently, it activates OmpR. Phosphorylated OmpR binds with high affinity to the regulatory regions of the ompF and ompC porin genes to regulate their transcription. We set out to visualize these two-components in single bacterial cells during different environmental stress conditions and to examine the subsequent modifications to the bacterial nucleoid as a result. We created a chromosomally-encoded, active, fluorescent OmpR-PAmCherry fusion protein and compared its expression levels with RNA polymerase. Quantitative western blotting had indicated that these two proteins were expressed at similar levels. From our images, it is evident that OmpR is significantly less abundant compared to RNA polymerase. In cross-sectional axial images, we observed OmpR molecules closely juxtaposed near the inner membrane during acidic and hyposomotic growth. In acidic conditions, the chromosome was compacted. Surprisingly, under acidic conditions, we also observed evidence of a spatial correlation between the DNA and the inner membrane, suggesting a mechanical link through an active DNA-OmpR-EnvZ complex. This work represents the first direct visualization of a response regulator with respect to the bacterial chromosome.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1297-1308 |
| Number of pages | 12 |
| Journal | Integrative Biology (United Kingdom) |
| Volume | 7 |
| Issue number | 10 |
| DOIs | |
| State | Published - Oct 1 2015 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Biophysics
- Biochemistry
Cite this
Single cell super-resolution imaging of E. coli OmpR during environmental stress. / Foo, Yong Hwee; Spahn, Christoph; Zhang, Hongfang; Heilemann, Mike; Kenney, Linda.
In: Integrative Biology (United Kingdom), Vol. 7, No. 10, 01.10.2015, p. 1297-1308.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Single cell super-resolution imaging of E. coli OmpR during environmental stress
AU - Foo, Yong Hwee
AU - Spahn, Christoph
AU - Zhang, Hongfang
AU - Heilemann, Mike
AU - Kenney, Linda
PY - 2015/10/1
Y1 - 2015/10/1
N2 - Two-component signaling systems are a major strategy employed by bacteria, and to some extent, yeast and plants, to respond to environmental stress. The EnvZ/OmpR system in E. coli responds to osmotic and acid stress and is responsible for regulating the protein composition of the outer membrane. EnvZ is a histidine kinase located in the inner membrane. Upon activation, it is autophosphorylated by ATP and subsequently, it activates OmpR. Phosphorylated OmpR binds with high affinity to the regulatory regions of the ompF and ompC porin genes to regulate their transcription. We set out to visualize these two-components in single bacterial cells during different environmental stress conditions and to examine the subsequent modifications to the bacterial nucleoid as a result. We created a chromosomally-encoded, active, fluorescent OmpR-PAmCherry fusion protein and compared its expression levels with RNA polymerase. Quantitative western blotting had indicated that these two proteins were expressed at similar levels. From our images, it is evident that OmpR is significantly less abundant compared to RNA polymerase. In cross-sectional axial images, we observed OmpR molecules closely juxtaposed near the inner membrane during acidic and hyposomotic growth. In acidic conditions, the chromosome was compacted. Surprisingly, under acidic conditions, we also observed evidence of a spatial correlation between the DNA and the inner membrane, suggesting a mechanical link through an active DNA-OmpR-EnvZ complex. This work represents the first direct visualization of a response regulator with respect to the bacterial chromosome.
AB - Two-component signaling systems are a major strategy employed by bacteria, and to some extent, yeast and plants, to respond to environmental stress. The EnvZ/OmpR system in E. coli responds to osmotic and acid stress and is responsible for regulating the protein composition of the outer membrane. EnvZ is a histidine kinase located in the inner membrane. Upon activation, it is autophosphorylated by ATP and subsequently, it activates OmpR. Phosphorylated OmpR binds with high affinity to the regulatory regions of the ompF and ompC porin genes to regulate their transcription. We set out to visualize these two-components in single bacterial cells during different environmental stress conditions and to examine the subsequent modifications to the bacterial nucleoid as a result. We created a chromosomally-encoded, active, fluorescent OmpR-PAmCherry fusion protein and compared its expression levels with RNA polymerase. Quantitative western blotting had indicated that these two proteins were expressed at similar levels. From our images, it is evident that OmpR is significantly less abundant compared to RNA polymerase. In cross-sectional axial images, we observed OmpR molecules closely juxtaposed near the inner membrane during acidic and hyposomotic growth. In acidic conditions, the chromosome was compacted. Surprisingly, under acidic conditions, we also observed evidence of a spatial correlation between the DNA and the inner membrane, suggesting a mechanical link through an active DNA-OmpR-EnvZ complex. This work represents the first direct visualization of a response regulator with respect to the bacterial chromosome.
UR - http://www.scopus.com/inward/record.url?scp=84943393630&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84943393630&partnerID=8YFLogxK
U2 - 10.1039/c5ib00077g
DO - 10.1039/c5ib00077g
M3 - Article
C2 - 26156621
AN - SCOPUS:84943393630
VL - 7
SP - 1297
EP - 1308
JO - Integrative Biology (United Kingdom)
JF - Integrative Biology (United Kingdom)
SN - 1757-9694
IS - 10
ER -