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@ARTICLE{Burgstaller:862520,
author = {Burgstaller, Sandra and Bischof, Helmut and Gensch, Thomas
and Stryeck, Sarah and Gottschalk, Benjamin and
Ramadani-Muja, Jeta and Eroglu, Emrah and Rost, Rene and
Balfanz, Sabine and Baumann, Arnd and Waldeck-Weiermair,
Markus and Hay, Jesse C. and Madl, Tobias and Graier,
Wolfgang F. and Malli, Roland},
title = {p{H}-{L}emon, a {F}luorescent {P}rotein-{B}ased p{H}
{R}eporter for {A}cidic {C}ompartments},
journal = {ACS sensors},
volume = {4},
number = {4},
issn = {2379-3694},
address = {Washington, DC},
publisher = {ACS Publications},
reportid = {FZJ-2019-02823},
pages = {883 - 891},
year = {2019},
abstract = {Distinct subcellular pH levels, especially in lysosomes and
endosomes, are essential for the degradation, modification,
sorting, accumulation, and secretion of macromolecules.
Here, we engineered a novel genetically encoded pH probe by
fusing the pH-stable cyan fluorescent protein (FP) variant,
mTurquoise2, to the highly pH-sensitive enhanced yellow
fluorescent protein, EYFP. This approach yielded a
ratiometric biosensor—referred to as pH-Lemon—optimized
for live imaging of distinct pH conditions within acidic
cellular compartments. Protonation of pH-Lemon under acidic
conditions significantly decreases the yellow fluorescence
while the cyan fluorescence increases due to reduced
Förster resonance energy transfer (FRET) efficiency.
Because of its freely reversible and ratiometric responses,
pH-Lemon represents a fluorescent biosensor for pH dynamics.
pH-Lemon also shows a sizable pH-dependent fluorescence
lifetime change that can be used in fluorescence lifetime
imaging microscopy as an alternative observation method for
the study of pH in acidic cellular compartments. Fusion of
pH-Lemon to the protein microtubule-associated protein
1A/1B-light chain 3B (LC3B), a specific marker of autophagic
membranes, resulted in its targeting within autolysosomes of
HeLa cells. Moreover, fusion of pH-Lemon to a
glycophosphatidylinositol (GPI) anchor allowed us to monitor
the entire luminal space of the secretory pathway and the
exoplasmic leaflet of the plasma membrane. Utilizing this
new pH probe, we revealed neutral and acidic vesicles and
substructures inside cells, highlighting compartments of
distinct pH throughout the endomembrane system. These data
demonstrate, that this novel pH sensor, pH-Lemon, is very
suitable for the study of local pH dynamics of subcellular
microstructures in living cells.},
cin = {ICS-4},
ddc = {570},
cid = {I:(DE-Juel1)ICS-4-20110106},
pnm = {552 - Engineering Cell Function (POF3-552)},
pid = {G:(DE-HGF)POF3-552},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:30864782},
UT = {WOS:000466442500013},
doi = {10.1021/acssensors.8b01599},
url = {https://juser.fz-juelich.de/record/862520},
}