A phosphatidylinositol (4,5)-bisphosphate binding site within mu2-adaptin regulates clathrin-mediated endocytosis.
The clathrin adaptor complex AP-2 serves to coordinate clathrin-coated pit assembly with the sorting of transmembrane cargo proteins at the plasmalemma. How precisely AP-2 assembly and cargo protein recognition at sites of endocytosis are regulated has remained unclear, but recent evidence implicates phosphoinositides, in particular phosphatidylinositol (4,5)-bisphosphate (PI[4,5]P2), in these processes. Here we have identified and functionally characterized a conserved binding site for PI(4,5)P2 within mu2-adaptin, the medium chain of the clathrin adaptor complex AP-2. Mutant mu2 lacking a cluster of conserved lysine residues fails to bind PI(4,5)P2 and to compete the recruitment of native clathrin/AP-2 to PI(4,5)P2-containing liposomes or to presynaptic membranes. Moreover, we show that expression of mutant mu2 inhibits receptor-mediated endocytosis in living cells. We suggest that PI(4,5)P2 binding to mu2-adaptin regulates clathrin-mediated endocytosis and thereby may contribute to structurally linking cargo recognition to coat formation.
Zentrum für Biochemie and Molekulare Zellbiologie, Department of Biochemistry II, University of Göttingen, Humboldtalle 23, D-37073 Göttingen, Germany.
Gundula
G
Rohde
Dirk
D
Wenzel
Volker
V
Haucke
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033963:Adaptor Protein Complex 3; D033983:Adaptor Protein Complex mu Subunits; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002966:Clathrin / Q000737:chemistry / Q000378:metabolism*; D006224:Cricetinae; D004705:Endocytosis; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats
D033961:Adaptor Protein Complex 1; D033962:Adaptor Protein Complex 2; D033963:Adaptor Protein Complex 3; D033983:Adaptor Protein Complex mu Subunits; D033942:Adaptor Proteins, Vesicular Transport; C510072:Ap3m2 protein, rat; D002352:Carrier Proteins; D002966:Clathrin; D008565:Membrane Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; C499065:adaptor protein complex 1, mu 2 subunit; C438886:adaptor protein complex 2, mu 2 subunit