title: NECAP 1 regulates AP-2 interactions to control vesicle size, number, and cargo during clathrin-mediated endocytosis.
abstract: AP-2 is the core-organizing element in clathrin-mediated endocytosis. During the formation of clathrin-coated vesicles, clathrin and endocytic accessory proteins interact with AP-2 in a temporally and spatially controlled manner, yet it remains elusive as to how these interactions are regulated. Here, we demonstrate that the endocytic protein NECAP 1, which binds to the α-ear of AP-2 through a C-terminal WxxF motif, uses an N-terminal PH-like domain to compete with clathrin for access to the AP-2 β2-linker, revealing a means to allow AP-2-mediated coordination of accessory protein recruitment and clathrin polymerization at sites of vesicle formation. Knockdown and functional rescue studies demonstrate that through these interactions, NECAP 1 and AP-2 cooperate to increase the probability of clathrin-coated vesicle formation and to control the number, size, and cargo content of the vesicles. Together, our data demonstrate that NECAP 1 modulates the AP-2 interactome and reveal a new layer of organizational control within the endocytic machinery.
authors.affiliation: Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada ; Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts, United States of America.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Ritter
authors.forename: Sebastian
authors.initials: S
authors.lastname: Murphy
authors.forename: Hatem
authors.initials: H
authors.lastname: Dokainish
authors.forename: Martine
authors.initials: M
authors.lastname: Girard
authors.forename: Manasa V
authors.initials: MV
authors.lastname: Gudheti
authors.forename: Guennadi
authors.initials: G
authors.lastname: Kozlov
authors.forename: Marilene
authors.initials: M
authors.lastname: Halin
authors.forename: Jacynthe
authors.initials: J
authors.lastname: Philie
authors.forename: Erik M
authors.initials: EM
authors.lastname: Jorgensen
authors.forename: Kalle
authors.initials: K
authors.lastname: Gehring
authors.forename: Peter S
authors.initials: PS
authors.lastname: McPherson
mesh_terms: D033962:Adaptor Protein Complex 2 / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D019556:COS Cells; D002522:Chlorocebus aethiops; D002966:Clathrin / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism*; D017124:Conserved Sequence; D004705:Endocytosis; D055785:Gene Knockdown Techniques; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D013572:Synaptic Vesicles / Q000378:metabolism
chemical_list: D033962:Adaptor Protein Complex 2; D002966:Clathrin; D008565:Membrane Proteins