Novel binding sites on clathrin and adaptors regulate distinct aspects of coat assembly.
Clathrin-coated vesicles (CCVs) sort proteins at the plasma membrane, endosomes and trans Golgi network for multiple membrane traffic pathways. Clathrin recruitment to membranes and its self-assembly into a polyhedral coat depends on adaptor molecules, which interact with membrane-associated vesicle cargo. To determine how adaptors induce clathrin recruitment and assembly, we mapped novel interaction sites between these coat components. A site in the ankle domain of the clathrin triskelion leg was identified that binds a common site on the appendages of tetrameric [AP1 and AP2] and monomeric (GGA1) adaptors. Mutagenesis and modeling studies suggested that the clathrin-GGA1 appendage interface is nonlinear, unlike other peptide-appendage interactions, but overlaps with a sandwich domain binding site for accessory protein peptides, allowing for competitive regulation of coated vesicle formation. A novel clathrin box in the GGA1 hinge region was also identified and shown to mediate membrane recruitment of clathrin, while disruption of the clathrin-GGA1 appendage interaction did not affect recruitment. Thus, the distinct sites for clathrin-adaptor interactions perform distinct functions, revealing new aspects to regulation of CCV formation.
Knuehl
Christine
C
The G.W. Hooper Foundation, Departments of Microbiology and Immunology, Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California, San Francisco, CA 94143-0552, USA.
Chen
Chih-Ying
CY
Manalo
Venus
V
Hwang
Peter K
PK
Ota
Nobuyuki
N
Brodsky
Frances M
FM
D033942:Adaptor Proteins, Vesicular Transport / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002966:Clathrin / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D041681:NIH 3T3 Cells; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary
D033942:Adaptor Proteins, Vesicular Transport; D002966:Clathrin