In non-neuronal cells, the stoichiometry of heavy chains to light chains might not always be one-to-one 18. This six protein complex is composed of three clathrin heavy chains and three smaller clathrin light chains (CLCs) that form a three-legged pinwheel (Supplementary Fig. The basic unit of the clathrin lattice is the clathrin triskelion 15, 16.
Determining the nanoscale localizations 4, 5, numbers 6, 7, accumulation dynamics 8, 9, interactions 1, and conformations 10, 11 of these proteins in cells is important for understanding how the endocytic machinery operates in health and disease.Īssembly and curvature of clathrin lattices are required steps to build cargo-loaded vesicles 12, 13, 14.
Over 50 proteins have been implicated in this process 2, 3. Thus, a specific conformational switch in the light chain regulates lattice curvature and endocytosis in mammalian cells.Ĭlathrin-mediated endocytosis is the primary internalization pathway in eukaryotic cells and is key to many processes, including nutrient uptake, excitability, signaling, and the recycling of membrane components 1. Preventing this conformational switch with chemical tools increases lattice sizes and inhibits endocytosis. We discover that the N-terminus of CLC repositions away from the plasma membrane and triskelia vertex as coats curve. With FRET-CLEM, we measure conformational changes in clathrin at thousands of individual morphologically distinct clathrin-coated structures. Here, we develop a new correlative fluorescence resonance energy transfer (FRET) and platinum replica electron microscopy method, named FRET-CLEM. Structural changes in CLC have been shown to regulate triskelia assembly in solution, yet the nature of these changes, and their effects on lattice growth, curvature, and endocytosis in cells are unknown. Three clathrin heavy chains associated with clathrin light chains (CLC) assemble into triskelia that link into a geometric lattice that curves to drive endocytosis. Conformational changes in endocytic proteins are regulators of clathrin-mediated endocytosis.
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