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Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells
Protein-protein interaction studies provide valuable insights into cellular signaling. Brassinosteroid (BR) signaling is initiated by the hormone-binding receptor Brassinosteroid Insensitive 1 (BRI1) and its co-receptor BRI1 Associated Kinase 1 (BAK1). BRI1 and BAK1 were shown to interact independen...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9571070/ https://www.ncbi.nlm.nih.gov/pubmed/36235497 http://dx.doi.org/10.3390/plants11192630 |
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author | Glöckner, Nina zur Oven-Krockhaus, Sven Rohr, Leander Wackenhut, Frank Burmeister, Moritz Wanke, Friederike Holzwart, Eleonore Meixner, Alfred J. Wolf, Sebastian Harter, Klaus |
author_facet | Glöckner, Nina zur Oven-Krockhaus, Sven Rohr, Leander Wackenhut, Frank Burmeister, Moritz Wanke, Friederike Holzwart, Eleonore Meixner, Alfred J. Wolf, Sebastian Harter, Klaus |
author_sort | Glöckner, Nina |
collection | PubMed |
description | Protein-protein interaction studies provide valuable insights into cellular signaling. Brassinosteroid (BR) signaling is initiated by the hormone-binding receptor Brassinosteroid Insensitive 1 (BRI1) and its co-receptor BRI1 Associated Kinase 1 (BAK1). BRI1 and BAK1 were shown to interact independently with the Receptor-Like Protein 44 (RLP44), which is implicated in BRI1/BAK1-dependent cell wall integrity perception. To demonstrate the proposed complex formation of BRI1, BAK1 and RLP44, we established three-fluorophore intensity-based spectral Förster resonance energy transfer (FRET) and FRET-fluorescence lifetime imaging microscopy (FLIM) for living plant cells. Our evidence indicates that RLP44, BRI1 and BAK1 form a ternary complex in a distinct plasma membrane nanodomain. In contrast, although the immune receptor Flagellin Sensing 2 (FLS2) also forms a heteromer with BAK1, the FLS2/BAK1 complexes are localized to other nanodomains. In conclusion, both three-fluorophore FRET approaches provide a feasible basis for studying the in vivo interaction and sub-compartmentalization of proteins in great detail. |
format | Online Article Text |
id | pubmed-9571070 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-95710702022-10-17 Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells Glöckner, Nina zur Oven-Krockhaus, Sven Rohr, Leander Wackenhut, Frank Burmeister, Moritz Wanke, Friederike Holzwart, Eleonore Meixner, Alfred J. Wolf, Sebastian Harter, Klaus Plants (Basel) Article Protein-protein interaction studies provide valuable insights into cellular signaling. Brassinosteroid (BR) signaling is initiated by the hormone-binding receptor Brassinosteroid Insensitive 1 (BRI1) and its co-receptor BRI1 Associated Kinase 1 (BAK1). BRI1 and BAK1 were shown to interact independently with the Receptor-Like Protein 44 (RLP44), which is implicated in BRI1/BAK1-dependent cell wall integrity perception. To demonstrate the proposed complex formation of BRI1, BAK1 and RLP44, we established three-fluorophore intensity-based spectral Förster resonance energy transfer (FRET) and FRET-fluorescence lifetime imaging microscopy (FLIM) for living plant cells. Our evidence indicates that RLP44, BRI1 and BAK1 form a ternary complex in a distinct plasma membrane nanodomain. In contrast, although the immune receptor Flagellin Sensing 2 (FLS2) also forms a heteromer with BAK1, the FLS2/BAK1 complexes are localized to other nanodomains. In conclusion, both three-fluorophore FRET approaches provide a feasible basis for studying the in vivo interaction and sub-compartmentalization of proteins in great detail. MDPI 2022-10-06 /pmc/articles/PMC9571070/ /pubmed/36235497 http://dx.doi.org/10.3390/plants11192630 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Glöckner, Nina zur Oven-Krockhaus, Sven Rohr, Leander Wackenhut, Frank Burmeister, Moritz Wanke, Friederike Holzwart, Eleonore Meixner, Alfred J. Wolf, Sebastian Harter, Klaus Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title | Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title_full | Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title_fullStr | Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title_full_unstemmed | Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title_short | Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells |
title_sort | three-fluorophore fret enables the analysis of ternary protein association in living plant cells |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9571070/ https://www.ncbi.nlm.nih.gov/pubmed/36235497 http://dx.doi.org/10.3390/plants11192630 |
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