Mapping Transient Protein Interactions at the Nanoscale in Living Mammalian Cells.
Authors of this article are:
De Keersmaecker H, Camacho R, Rantasa DM, Fron E, Uji-I H, Mizuno H, Rocha S.
A summary of the article is shown below:
Protein-protein interactions (PPIs) form the basis of cellular processes, regulating cell behavior and fate. PPIs can be extremely transient in nature, which hinders their detection. In addition, traditional biochemical methods provided limited information on the spatial distribution and temporal dynamics of PPIs that is crucial for their regulation in the crowded cellular environment. Given the pivotal role of membrane micro- and nanodomains in the regulation of PPIs at the plasma membrane, the development of methods to visualize PPIs with a high spatial resolution is imperative. Here, we present a super-resolution fluorescence microscopy technique that can detect and map short-lived transient protein-protein interactions on a nanometer scale in the cellular environment. This imaging method is based on single-molecule fluorescence microscopy and exploits the effect of the difference in the mobility between cytosolic and membrane-bound proteins in the recorded fluorescence signals. After the development of the proof of concept using a model system based on membrane-bound modular protein domains and fluorescently labeled peptides, we applied this imaging approach to investigate the interactions of cytosolic proteins involved in the epidermal growth factor signaling pathway (namely, Grb2, c-Raf, and PLCγ1). The detected clusters of Grb2 and c-Raf were correlated with the distribution of the receptor at the plasma membrane. Additionally, the interactions of wild type PLCγ1 were compared with those detected with truncated mutants, which provided important information regarding the role played by specific domains in the interaction with the membrane. The results presented here demonstrate the potential of this technique to unravel the role of membrane heterogeneity in the spatiotemporal regulation of cell signaling.
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This article is a good source of information and a good way to become familiar with topics such as:
cell signaling;epidermal growth factor;fluorescence microscopy;interaction map;live-cell imaging;protein−protein interactions;single-molecule localization microscopy
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