Cryo–light microscopy with angstrom precision deciphers structural conformations of PIEZO1 in its native state

Investigations based on cryo–electron microscopy (cryo-EM), atomic force microscopy, and super-resolution microscopy reveal a symmetric trimer with propeller-like blades for the mechanosensitive ion channel PIEZO. However, a conclusive understanding of its conformations in the cell membrane is lacking. Here, we implement a high-vacuum cryogenic shuttle to transfer shock-frozen cell membranes in and out of a cryostat designed for single-particle cryo–light microscopy (spCryo-LM). By localizing fluorescent labels placed at the extremities of the blades of the mouse PIEZO1 protein in unroofed cell membranes, we ascertain three configurations with radii of 6, 12, and 20 nanometers as projected onto the membrane plane. We elaborate on the correspondence of these data with previous reports in the literature. The combination of spCryo-LM with cryo-EM promises to provide quantitative insights into the structure and function of biomolecular complexes in their native environments without the need for chemical fixation or protein isolation, ushering in a new regime of correlative studies in structural biology.