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@ARTICLE{Fischer:614229,
author = {Fischer, Alexandra A. M. and Schatz, Larissa and Baaske,
Julia and Römer, Winfried and Weber, Wilfried and
Thünauer, Roland},
title = {{R}eal‐time monitoring of cell surface protein arrival
with split luciferases},
journal = {Traffic},
volume = {24},
number = {10},
issn = {1398-9219},
address = {Oxford},
publisher = {Wiley-Blackwell},
reportid = {PUBDB-2024-05769},
pages = {453 - 462},
year = {2023},
abstract = {Each cell in a multicellular organism permanently adjusts
the concentration of its cell surface proteins. In
particular, epithelial cells tightly control the number of
carriers, transporters and cell adhesion proteins at their
plasma membrane. However, sensitively measuring the cell
surface concentration of a particular protein of interest in
live cells and in real time represents a considerable
challenge. Here, we introduce a novel approach based on
split luciferases, which uses one luciferase fragment as a
tag on the protein of interest and the second fragment as a
supplement to the extracellular medium. Once the protein of
interest arrives at the cell surface, the luciferase
fragments complement and generate luminescence. We compared
the performance of split Gaussia luciferase and split
Nanoluciferase by using a system to synchronize biosynthetic
trafficking with conditional aggregation domains. The best
results were achieved with split Nanoluciferase, for which
luminescence increased more than 6000-fold upon
recombination. Furthermore, we showed that our approach can
separately detect and quantify the arrival of membrane
proteins at the apical and basolateral plasma membrane in
single polarized epithelial cells by detecting the
luminescence signals with a microscope, thus opening novel
avenues for characterizing the variations in trafficking in
individual epithelial cells.},
cin = {CSSB-CF-ALFM},
ddc = {570},
cid = {I:(DE-H253)CSSB-CF-ALFM-20210629},
pnm = {899 - ohne Topic (POF4-899) / DFG project
G:(GEPRIS)390939984 - EXC 2189: CIBSS - Centre for
Integrative Biological Signalling Studies (390939984)},
pid = {G:(DE-HGF)POF4-899 / G:(GEPRIS)390939984},
experiment = {EXP:(DE-MLZ)NOSPEC-20140101},
typ = {PUB:(DE-HGF)16},
pubmed = {pmid:37403269},
UT = {WOS:001024226500001},
doi = {10.1111/tra.12908},
url = {https://bib-pubdb1.desy.de/record/614229},
}