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Single cell super-resolution imaging of E. coli OmpR during environmental stress.
journal contributionposted on 2016-08-02, 00:00 authored by Foo YH, Spahn C, Zhang H, Heilemann M, Kenney LJ
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.
MH and CKS acknowledge funding by the German Science Foundation (DFG, grant EXC 115) and by Goethe University Frankfurt. LJK received support from Research Centre of Excellence Grant in Mechanobiology from the Ministry of Education, Singapore and VA 5IOBX000372.
Publisher StatementThis is a copy of an article published in Integrative Biology. © 2015 Royal Society of Chemistry Publications.
PublisherRoyal Society of Chemistry