Real-time imaging of the intracellular glutathione redox potential
Abstract
Dynamic analysis of redox-based processes in living cells is now restricted by the lack of appropriate redox biosensors. Conventional redox-sensitive GFPs (roGFPs) are limited by undefined specificity and slow response to changes in redox potential. In this study we demonstrate that the fusion of human glutaredoxin-1 (Grx1) to roGFP2 facilitates specific real-time equilibration between the sensor protein and the glutathione redox couple. The Grx1-roGFP2 fusion protein allowed dynamic live imaging of the glutathione redox potential (E(GSH)) in different cellular compartments with high sensitivity and temporal resolution. The biosensor detected nanomolar changes in oxidized glutathione (GSSG) against a backdrop of millimolar reduced glutathione (GSH) on a scale of seconds to minutes. It facilitated the observation of redox changes associated with growth factor availability, cell density, mitochondrial depolarization, respiratory burst activity and immune receptor stimulation.
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This recombinant sensor apparently equilibrates more quickly with cellular GSSG than redox-sensitive probes that lack the Grx domain in cis with the redox-sensor domain.
“This recombinant sensor apparently equilibrates more quickly with cellular GSSG than redox-sensitive probes that lack the Grx domain in cis with the redox-sensor domain.”
From Redox regulation of mitochondrial function (2012) · cites this paper as [163]
A similar biodetector, engineered by combining human Grx with a redox-sensitive GFP, was used to image GSSG-redox changes within human cells.
“A similar biodetector, engineered by combining human Grx with a redox-sensitive GFP, was used to image GSSG-redox changes within human cells (163).”
From Redox regulation of mitochondrial function (2012) · cites this paper as [163]
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- Authors
- Gutscher, M., Pauleau, A.-L., Marty, L., Brach, T., Wabnitz, G. H., Samstag, Y., Meyer, A. J., & Dick, T. P.
- Affiliations
- German Cancer Research Center (DE)
- Heidelberg University (DE)
- Corresponding author
- Dick, Tobias P
- ORCID
- Venue
- Nature Methods
- Published
- 2008-05-11
- Volume
- 5
- Issue
- 6
- Pages
- 553-559
- ISSN
- 1548-7091 (ISSN-L)
- 1548-7105
- OpenAlex
- W2100811292
- Cited by
- 894
- References
- 37
- Citations by year
- 2026: 38 · 2025: 51 · 2024: 70 · 2023: 43 · 2022: 72 · 2021: 53 · 2020: 58 · 2019: 67 · 2018: 62 · 2017: 67 · 2016: 50 · 2015: 54 · 2014: 48 · 2013: 45 · 2012: 30
- FWCI
- 11.78
- Type
- journal article
- Language
- en
- Open access
- closed
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Methods (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Redox biology and oxidative stress
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biochemistry › Sulfur Compounds in Biology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Keywords
- glutathione redox potential, glutaredoxin 1, roGFP2, ratiometric biosensor, glutathione disulfide, reduced glutathione, mitochondrial depolarization, respiratory burst, cell density, growth factor signaling
- MeSH terms
- Biotechnology, Disulfides, Glutathione, HeLa Cells, Humans, Mitochondria, Oxidation-Reduction, Oxygen, Recombinant Fusion Proteins, Sensitivity and Specificity, Thioredoxins, Biosensing Techniques, Apoptosis, Green Fluorescent Proteins, Glutaredoxins
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