[HTML][HTML] A genetically encoded single-wavelength sensor for imaging cytosolic and cell surface ATP

MA Lobas, R Tao, J Nagai, MT Kronschläger… - Nature …, 2019 - nature.com
MA Lobas, R Tao, J Nagai, MT Kronschläger, PM Borden, JS Marvin, LL Looger, BS Khakh
Nature communications, 2019nature.com
Abstract Adenosine 5′ triphosphate (ATP) is a universal intracellular energy source and an
evolutionarily ancient, ubiquitous extracellular signal in diverse species. Here, we report the
generation and characterization of single-wavelength genetically encoded fluorescent
sensors (iATPSnFRs) for imaging extracellular and cytosolic ATP from insertion of circularly
permuted superfolder GFP into the epsilon subunit of F0F1-ATPase from Bacillus PS3. On
the cell surface and within the cytosol, iATPSnFR1. 0 responds to relevant ATP …
Abstract
Adenosine 5′ triphosphate (ATP) is a universal intracellular energy source and an evolutionarily ancient, ubiquitous extracellular signal in diverse species. Here, we report the generation and characterization of single-wavelength genetically encoded fluorescent sensors (iATPSnFRs) for imaging extracellular and cytosolic ATP from insertion of circularly permuted superfolder GFP into the epsilon subunit of F0F1-ATPase from Bacillus PS3. On the cell surface and within the cytosol, iATPSnFR1.0 responds to relevant ATP concentrations (30 μM to 3 mM) with fast increases in fluorescence. iATPSnFRs can be genetically targeted to specific cell types and sub-cellular compartments, imaged with standard light microscopes, do not respond to other nucleotides and nucleosides, and when fused with a red fluorescent protein function as ratiometric indicators. After careful consideration of their modest pH sensitivity, iATPSnFRs represent promising reagents for imaging ATP in the extracellular space and within cells during a variety of settings, and for further application-specific refinements.
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