Journal Article

PUBDB-2024-05769

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Real‐time monitoring of cell surface protein arrival with split luciferases

Fischer, A. A. M. ; Schatz, L.CSSB* ; Baaske, J. ; Römer, W. ; Weber, W. ; Thünauer, R. (Corresponding author)Extern*CSSB*

2023
Wiley-Blackwell Oxford

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Please use a persistent id in citations: doi:10.1111/tra.12908  doi:10.3204/PUBDB-2024-05769

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.

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Contributing Institute(s):

1. CSSB-CF-ALFM (CSSB-CF-ALFM)

Research Program(s):

1. 899 - ohne Topic (POF4-899) (POF4-899)
2. DFG project G:(GEPRIS)390939984 - EXC 2189: CIBSS - Centre for Integrative Biological Signalling Studies (390939984) (390939984)

Experiment(s):

1. No specific instrument

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Database coverage:
; ; ; BIOSIS Previews ; Biological Abstracts ; Clarivate Analytics Master Journal List ; Current Contents - Life Sciences ; DEAL Wiley ; Ebsco Academic Search ; Essential Science Indicators ; IF < 5 ; JCR ; SCOPUS ; Science Citation Index Expanded ; Web of Science Core Collection

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