Sustained accurate recording of intracellular acidification in living tissues with a photo-controllable bioluminescent protein

Regulation of an intracellular acidic environment plays a pivotal role in biological processes and functions. However, spatiotemporal analysis of the acidification in complex tissues of living subjects persists as an important challenge. We developed a photo-inactivatable bioluminescent indicator, b...

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Published inProceedings of the National Academy of Sciences - PNAS Vol. 110; no. 23; pp. 9332 - 9337
Main Authors Hattori, Mitsuru, Haga, Sanae, Takakura, Hideo, Ozaki, Michitaka, Ozawa, Takeaki
Format Journal Article
LanguageEnglish
Published United States National Academy of Sciences 04.06.2013
National Acad Sciences
Subjects
Acid-Base Equilibrium - physiology
Acidification
Adenosine triphosphatase
Animals
Apoptosis
Arithmetic mean
Avena - metabolism
Biological Sciences
Bioluminescence
Fluorescence
HEK293 cells
Hydrogen-Ion Concentration
Imaging
Indicators and Reagents - chemistry
Irradiation
Ischemia
Luminescent Measurements - methods
Mice
Microscopes
Monitoring, Physiologic - methods
Organelles
Oxygen - metabolism
Photic Stimulation
Phototropins - metabolism
Phototropins - physiology
Physical Sciences
Protein Structure, Tertiary - physiology
Proteins
Rodents
Tissues
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Summary:Regulation of an intracellular acidic environment plays a pivotal role in biological processes and functions. However, spatiotemporal analysis of the acidification in complex tissues of living subjects persists as an important challenge. We developed a photo-inactivatable bioluminescent indicator, based on a combination of luciferase-fragment complementation and a photoreaction of a light, oxygen, and voltage domain from Avena sativa Phototropin1 (LOV2), to visualize temporally dynamic acidification in living tissue samples. Bioluminescence of the indicator diminished upon light irradiation and it recovered gradually in the dark state thereafter. The recovery rate was remarkably sensitive to pH changes but unsusceptible to fluctuation of luciferin or ATP concentrations. Bioluminescence imaging, taken as an index of the recovery rates, enabled long-time recording of acidification in apoptotic and autophagous processes in a cell population and an ischemic condition in living mice. This technology using the indicator is widely applicable to sense organelle-specific acidic changes in target biological tissues.
Bibliography:http://dx.doi.org/10.1073/pnas.1304056110
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Edited by J. Woodland Hastings, Harvard University, Cambridge, MA, and approved April 23, 2013 (received for review March 12, 2013)
Author contributions: M.H., S.H., and T.O. designed research; M.H., S.H., H.T., and M.O. performed research; M.H., H.T., M.O., and T.O. analyzed data; and M.H. and T.O. wrote the paper.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.1304056110