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Tickets to ride: selecting cargo for clathrin-regulated internalization

Nature Reviews Molecular Cell Biology volume 10, pages 583–596 (2009)Cite this article

Key Points

  • Endocytosis dynamically regulates the composition of the plasma membrane in eukaryotic cells by moving transmembrane proteins, bound ligands and membrane lipids off the surface. Clathrin-coated vesicles are a major type of carrier for the transport of internalized cargoes, although there are several other clathrin-independent uptake pathways that operate in parallel at the cell surface.

  • Clathrin-mediated uptake of surface transmembrane cargo proteins can be constitutive or temporally regulated, depending on ligand binding or activation.

  • Packaging into clathrin-coated vesicles forming at the cell surface requires a positive sorting signal in the cytosolic domain of the cargo molecule. Regulated endocytosis often involves addition of a reversible post-translational sorting tag.

  • There is an array of structurally distinct sorting signals that are decoded by different cytosolic adaptor proteins. The heterotetrameric AP-2 adaptor complex recognizes some types of cargo, whereas monomeric clathrin-associated sorting proteins (CLASPs) sort other signals into assembling vesicles.

  • CLASPs typically bind to lipids and to AP-2 and clathrin, which operate as organizational interaction nodes during vesicle assembly and budding. It is possible that segregated cargo-selective clathrin-coated structures might assemble at the cell surface, sorting transmembrane cargo into the cell in parallel but with different endosomal fates.

Abstract

Clathrin-mediated endocytosis oversees the constitutive packaging of selected membrane cargoes into transport vesicles that fuse with early endosomes. The process is responsive to activation of signalling receptors and ion channels, promptly clearing post-translationally tagged forms of cargo off the plasma membrane. To accommodate the diverse array of transmembrane proteins that are variably gathered into forming vesicles, a dedicated sorting machinery cooperates to ensure that non-competitive uptake from the cell surface occurs within minutes. Recent structural and functional data reveal remarkable plasticity in how disparate sorting signals are recognized by cargo-selective clathrin adaptors, such as AP-2. Cargo loading also seems to govern whether coats ultimately bud or dismantle abortively at the cell surface.

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Figure 1: Modular sorting codes.
Figure 2: Structural basis for cargo recognition.
Figure 3: Clathrin-coated structures at the cell surface.
Figure 4: The endocytic clathrin sorting interactome.

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Acknowledgements

Space constraints have unfortunately precluded discussion of many excellent and important studies in this area. I am grateful to N. Johnson, D. Owen and the anonymous referees for critical comments on the manuscript, to J. Heuser, R. Roth, S. Watkins and J. Thieman for beautiful microscopic images and to members of my laboratory for discussions. Support was from the US National Institutes of Health (R01 DK53249) and the American Heart Association (Award 0540007N).

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  1. Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, USA, 15261, Pennsylvania

    Linton M. Traub

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DATABASES

Protein DataBank

 1BXX

1KY7

1M7E

1W80

1YX6

2G30

2IV9

2JKR

2PR9

2VGL

2VX8

FURTHER INFORMATION

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Glossary

Clathrin

A triskelion-shaped protomer, composed of three heavy and three light chains, that polymerizes into a characteristic polyhedrally structured coat that is found on several intracellular membrane surfaces.

SNARE

(Soluble N-ethylmaleimide-sensitive factor attachment protein receptor). A member of a family of transmembrane proteins that assemble into heteromeric four-helix-bundle complexes to fuse membrane bilayers together.

Dynamin

A large regulatory GTPase that drives membrane scission when it is polymerized into a spiral at the base of deeply invaginated buds.

AAK1

(Adaptor-associated kinase 1). A Ser/Thr protein kinase that regulates endocytosis and cargo recognition.

Type I transmembrane protein

A single-pass transmembrane protein oriented with an extracellular N terminus and a cytosolic C terminus.

Type II transmembrane protein

A single-pass transmembrane protein oriented with an extracellular C terminus and a cytosolic N terminus.

AP-1

An AP-2-related heterotetrameric sorting adaptor that couples cargo selection with clathrin assembly at the TGN and/or in endosomes.

AP-3

An AP-2-related heterotetrameric sorting adaptor that is involved in sorting transmembrane cargoes through the endosomal system.

Integrin

A heterodimeric surface receptor that binds the extracellular matrix and couples to the actin cytoskeleton at the internal membrane surface.

FERM domain

A tripartite domain found in talin and ERM proteins that binds to integrins, thereby activating their extracellular domain through conformational rearrangements.

Focal adhesion

A large localized contact between a cell and the extracellular matrix. It is established through integrins and serves as an intracellular signalling station to arrange actin into stress fibres.

NUMB

A PTB domain-containing CLASP, first identified in D. melanogaster, that is involved in asymmetric positioning of cell surface receptors.

Fibronectin

A secreted extracellular matrix component that assembles into polymers on binding to integrins.

UBA domain

(Ubiquitin-associated domain). A small three-helix fold that binds to ubiquitin monomers or ubiquitin chains.

ENTH domain

(Epsin N-terminal homology domain). An all-α-helical module that binds to PtdIns(4,5)P2 and might change the properties of the underlying bilayer.

NPF motif

An Asn-Pro-Phe tripeptide sequence that binds to EH domains and is often tandemly arrayed in disordered polypeptide regions.

EH domain

(EPS15 homology domain). An ∼100-residue folded domain found in proteins that are involved in endocytosis and vesicular transport.

E3 ubiquitin ligase

An enzyme that catalyses the transfer of a thioester-linked reactive ubiquitin to an acceptor Lys residue on the substrate protein and determines substrate specificity.

E2 enzyme

The second intermediate in the three-step process of ubiquitylation. It delivers activated ubiquitin to an E3 ligase.

Deubiquitylating enzyme

An isopeptidase that removes ubiquitin monomers from acceptor proteins or assembled ubiquitin chains, thereby replenishing the pool of ubiquitin and allowing further ubiquitylation.

26S proteasome

A large proteolytic complex that generally degrades polyubiquitylated proteins in the cytosol.

WW domain

A small modular domain containing two crucial Trp residues that engage short Pro-rich interaction peptides with micromolar affinity to mediate protein–protein interactions.

G protein-coupled receptor

(GPCR). A member of the largest class of signalling receptors, which are defined by seven transmembrane α-helices. GPCRs couple a wide range of stimuli, including light, odorants, hormones and small bioactive molecules, to various intracellular signal transduction pathways.

Heterotrimeric G protein

A member of a family of membrane-apposed effectors of GPCRs that are composed of α, β and γ subunits and stimulate adenylyl cyclase, phospholipase C, ion channels or several other signalling molecules.

Longin domain

A modular domain found in several functionally discrete proteins that are involved in protein trafficking.

Retromer

A non-clathrin-based sorting coat that is assembled on endosomal tubules.

COPII

(Coat protein II). A complex that is involved in packaging cargoes into transport vesicles leaving the ER from discrete exit sites.

Delta

A transmembrane ligand for the Notch receptor that internalizes, along with the extracellular domain of Notch, after proteolysis triggered by Delta engagement.

Notch signalling

A conserved pathway in which proteolysis of the ligand-bound Notch receptor releases the intracytoplasmic domain, which enters the nucleus and regulates gene expression.

Imaginal discs

Sheets of epithelial cells in D. melanogaster larvae that, during pupation, develop into different adult body parts and structures.

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Traub, L. Tickets to ride: selecting cargo for clathrin-regulated internalization. Nat Rev Mol Cell Biol 10, 583–596 (2009). https://doi.org/10.1038/nrm2751

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