Non-invasive, ratiometric determination of intracellular pH in Pseudomonas species using a novel genetically encoded indicator.
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Your vote was cast
Thank you for your feedback
Thank you for your feedback
MetadataShow full item record
AbstractThe ability of Pseudomonas species to thrive in all major natural environments (i.e. terrestrial, freshwater and marine) is based on its exceptional capability to adapt to physicochemical changes. Thus, environmental bacteria have to tightly control the maintenance of numerous physiological traits across different conditions. The intracellular pH (pHi ) homoeostasis is a particularly important feature, since the pHi influences a large portion of the biochemical processes in the cell. Despite its importance, relatively few reliable, easy-to-implement tools have been designed for quantifying in vivo pHi changes in Gram-negative bacteria with minimal manipulations. Here we describe a convenient, non-invasive protocol for the quantification of the pHi in bacteria, which is based on the ratiometric fluorescent indicator protein PHP (pH indicator for Pseudomonas). The DNA sequence encoding PHP was thoroughly adapted to guarantee optimal transcription and translation of the indicator in Pseudomonas species. Our PHP-based quantification method demonstrated that pHi is tightly regulated over a narrow range of pH values not only in Pseudomonas, but also in other Gram-negative bacterial species such as Escherichia coli. The maintenance of the cytoplasmic pH homoeostasis in vivo could also be observed upon internal (e.g. redirection of glucose consumption pathways in P. putida) and external (e.g. antibiotic exposure in P. aeruginosa) perturbations, and the PHP indicator was also used to follow dynamic changes in the pHi upon external pH shifts. In summary, our work describes a reliable method for measuring pHi in Pseudomonas, allowing for the detailed investigation of bacterial pHi homoeostasis and its regulation.
CitationMicrob Biotechnol. 2019 Jul;12(4):799-813. doi: 10.1111/1751-7915.13439. Epub. 2019 Jun 4.
AffiliationHZI, Helmholtz -Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
The following license files are associated with this item:
Except where otherwise noted, this item's license is described as Attribution-NonCommercial-ShareAlike 4.0 International
- Optical Quantification of Intracellular pH in Drosophila melanogaster Malpighian Tubule Epithelia with a Fluorescent Genetically-encoded pH Indicator.
- Authors: Rossano AJ, Romero MF
- Issue date: 2017 Aug 11
- Intracellular pH imaging in cancer cells in vitro and tumors in vivo using the new genetically encoded sensor SypHer2.
- Authors: Shirmanova MV, Druzhkova IN, Lukina MM, Matlashov ME, Belousov VV, Snopova LB, Prodanetz NN, Dudenkova VV, Lukyanov SA, Zagaynova EV
- Issue date: 2015 Sep
- Imaging of Intracellular pH in Tumor Spheroids Using Genetically Encoded Sensor SypHer2.
- Authors: Zagaynova EV, Druzhkova IN, Mishina NM, Ignatova NI, Dudenkova VV, Shirmanova MV
- Issue date: 2017
- Intracellular pH Response to Weak Acid Stress in Individual Vegetative Bacillus subtilis Cells.
- Authors: Pandey R, Vischer NO, Smelt JP, van Beilen JW, Ter Beek A, De Vos WH, Brul S, Manders EM
- Issue date: 2016 Nov 1
- In vivo determination of organellar pH using a universal wavelength-based confocal microscopy approach.
- Authors: Pineda Rodó A, Váchová L, Palková Z
- Issue date: 2012