Work

Super‐Resolution Live‐Cell Mapping of Protein–Protein Interactions Using Chemogenetic Split Reporters and Stimulated Emission Depletion Microscopy

Year: 2025

Type: article

Abstract: The ability to map protein-protein interactions (PPI) within living cells at high spatial resolution is essential for unraveling their roles in cellular biology. Although several super-resolution microscopy techniques are available, visualizing PPI below the diffraction limit of light across entire live cells remains a challenge. An approach is introduced that combines the chemogenetic split fluorescent reporter splitFAST2 with stimulated emission depletion (STED) microscopy for sub-diffraction imaging of PPI. As the fluorescence of splitFAST2 is activated only where the two proteins interact, this system allows for highly precise and unambiguous localization in live cells, enhanced further by the rapid super-resolution imaging capabilities of STED. The improved spatial resolution of the approach enabled us to precisely map the subcellular localization of inducible interactions or constitutive interactions. Beyond simply detecting PPIs below the diffraction limit in whole living cells, splitFAST2 also serves as the basis for fluorescent probes with very low background, enabling effective subdiffraction imaging of repetitive cellular structures like filamentous actin and microtubules.

Source: ChemBioChem

DOI: 10.1002/cbic.202500754

Authors: Stephanie Board, Arnaud Gautier

Institutions: Centre National de la Recherche Scientifique, Université Paris Sciences et Lettres, Sorbonne Université, Chimie Physique et Chimie du Vivant, Institut Universitaire de France

Language: English

Topic: Advanced Fluorescence Microscopy Techniques

Subfield: Biophysics

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: hybrid

Estimated APC paid: $4,430