The Structures of Natively Assembled Clathrin Coated Vesicles

Abstract

Vesicle trafficking by clathrin coated vesicles (CCVs) is one of the major mechanisms by which proteins and nutrients are absorbed by the cell and transported between organelles. The individual proteins comprising the coated vesicles include clathrin heavy chain, clathrin light chain, and a variety of adaptor protein complexes. Much is known about the structures of the individual CCV components, but data are lacking about the structures of the fully assembled complexes together with membrane and in complex with cargo. Here we determined the structures of natively assembled CCVs in a variety of geometries. We show that the adaptor β2appendages crosslink adjacent CHC β-propellers and that the appendage densities reside almost exclusively in CCV hexagonal faces. We resolve how AP2 and other associated factors in hexagonal faces form an assembly hub with an extensive web of interactions between neighboring β-propellers and propose a structural model that explains how adaptor binding can direct the formation of pentagonal and hexagonal faces.

Multi‐modal adaptor‐clathrin contacts drive coated vesicle assembly

“…In comparing our results with those of Kovtun et al and Paraan et al, we see that differing sample conditions such as inclusion of membrane and cargo (Kovtun et al, 2020) or use of coated vesicles purified from source (Paraan et al, 2020) do not explain differences in b-appendage binding location since in our study with only two purified protein components we see multiple binding locations that either confirm (Kovtun et al, 2020) or are similar to (Paraan et al, 2020) those seen in more complex systems. One interpretation of this is that the long linker domain on b2-adaptin permits the bappendage to move freely, even within a coated vesicle, and form stable interactions unencumbered by the need for additional components or a specific orientation.…”

“…In contrast, Paraan et al isolated native coated vesicles from bovine brain and obtained a structure using single particle analysis. They observed density consistent with the b2-appendage, however it was in a different location, between two adjacent terminal domains (Paraan et al, 2020).…”

Clathrin packets move in slow axonal transport and deliver functional payloads to synapses

“…The structure of an axonal transport packet resembles clathrin-coated vesicles. However, coated vesicles are thought to be transient intermediates that deliver endocytosed vesicles and their contents either to Golgi/endosomes or back to the plasma membrane (for review, see Paraan et al, 2020), while the transport packets are long lasting structures that are (F) Quantification of fluorescence change after single-packet photoconversion in boutons. Schematic on left shows general principle of quantifying photoconverted and non-photoconverted (dark) regions; see STAR methods for details.…”

A conformational switch in clathrin light chain regulates lattice structure and endocytosis at the plasma membrane of mammalian cells

“…To test these models, we used FRET-CLEM. Specifically, the distances between the Nterminus and C-terminus of surrounding CLCs differ substantially between the extended and bent conformations according to structural models 17,45 . This substantial increase did not occur when expressing only EGFP-CLC.…”