Cryo-EM of multiple cage architectures reveals a universal mode of clathrin self-assembly

Abstract

Clathrin forms diverse lattice and cage structures that change size and shape rapidly in response to the needs of eukaryotic cells during clathrin-mediated endocytosis and intracellular trafficking. We present the cryo-EM structure and molecular model of assembled porcine clathrin, providing new insights into interactions that stabilise key elements of the clathrin lattice, namely, between Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

The Structures of Natively Assembled Clathrin Coated Vesicles

“…In this rigid-body fit, the edge of the vertex that is between two pentagonal faces exhibits an accurate fit, while the two other edges with one hexagonal face do not enclose the model starting at positions close to the knee domain. This discrepancy is due to our map deriving from the asymmetric vertex while theirs is from the symmetric one and is consistent with observations that the knee domain changes conformation to make hexagonal and pentagonal faces (14) while the proximal domain is stabilized by CLC. Using the asymmetric vertex structure from the mini-coat, a homology model of the bovine CHC and CLC was built and refined to our map ( Fig.…”

“…1A). The mini-coat, football, tennis ball, and barrel structures match four of the cage geometries that were recently reported by Morris et al (14) from in vitro reconstituted clathrin cages where they used a library of pre-generated cage geometries to sort their particles into the representative cage structures. This indicates that the cellular processes that assemble CCVs are similar to clathrin cages generated by reconstitution.…”

“…This interaction is similar to what was reported by Morris et al, but with small differences in the atomic model. Our model contains a small helix that is modelled as a loop in the previously published PDB-6SCT (14). Our change orients a number of hydrophobic residues to be packed against the trimerization domain ( Fig.…”

“…Clathrin together with the adaptors AP1 and AP2 act as assembly hubs due to their multiple binding sites for other adaptor/accessory proteins (15)(16)(17). The dynamic assembly of clathrin-coated vesicles has been investigated by super-resolution time-resolved fluorescence microscopy (8,11,18,19), and the self-assembly properties of clathrin have been understood by cryo-EM (14,20,21). However, it remains to be explained how clathrin assembly progresses into a closed fullerene-like cage with tight restraints on the configuration of hexagonal and pentagonal faces (22), in a mechanism that serves to transport cargo in lipid vesicles irrespective of cage geometry.…”

“…Both of these functions have been shown to be facilitated by the action of adaptor/accessory proteins which are often multi-functional and share overlapping functions (7)(8)(9). Though clathrin binding has been shown to be dispensable for curvature generation, it plays a major role in organizing adaptors and cargo into spherical vesicles (10-12) by making geometrical cages of different sizes both in vitro and in vivo (13,14). Clathrin together with the adaptors AP1 and AP2 act as assembly hubs due to their multiple binding sites for other adaptor/accessory proteins (15)(16)(17).…”