4polar-STORM polarized super-resolution imaging of actin filament organization in cells

Caio Vaz Rimoli, Cesar Augusto Valades-Cruz, Valentina Curcio, Manos Mavrakis () and Sophie Brasselet ()
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Caio Vaz Rimoli: Aix Marseille Univ, CNRS, Centrale Marseille, Institut Fresnel
Cesar Augusto Valades-Cruz: Institut Curie, PSL Research University, UMR144 CNRS, Space-Time imaging of organelles and Endomembranes Dynamics Team
Valentina Curcio: Aix Marseille Univ, CNRS, Centrale Marseille, Institut Fresnel
Manos Mavrakis: Aix Marseille Univ, CNRS, Centrale Marseille, Institut Fresnel
Sophie Brasselet: Aix Marseille Univ, CNRS, Centrale Marseille, Institut Fresnel

Nature Communications, 2022, vol. 13, issue 1, 1-13

Abstract: Abstract Single-molecule localization microscopy provides insights into the nanometer-scale spatial organization of proteins in cells, however it does not provide information on their conformation and orientation, which are key functional signatures. Detecting single molecules’ orientation in addition to their localization in cells is still a challenging task, in particular in dense cell samples. Here, we present a polarization-splitting scheme which combines Stochastic Optical Reconstruction Microscopy (STORM) with single molecule 2D orientation and wobbling measurements, without requiring a strong deformation of the imaged point spread function. This method called 4polar-STORM allows, thanks to a control of its detection numerical aperture, to determine both single molecules’ localization and orientation in 2D and to infer their 3D orientation. 4polar-STORM is compatible with relatively high densities of diffraction-limited spots in an image, and is thus ideally placed for the investigation of dense protein assemblies in cells. We demonstrate the potential of this method in dense actin filament organizations driving cell adhesion and motility.

Date: 2022
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DOI: 10.1038/s41467-022-27966-w

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