This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Iannolo, G. Articles by Keen, J. H. Search for Related Content PubMed PubMed Citation Articles by Iannolo, G. Articles by Keen, J. H.

Mapping of the Molecular Determinants Involved in the Interaction between eps15 and AP-2¹

Department of Experimental Oncology, European Institute of Oncology, Milan-20141, Italy [G. I., A. E. S., P. G. P., P. P. D. F.]; Istituto di Patologia Medica, Facolta' di Medicina e Chirurgia, Parma-43100, Italy [P. G. P.]; Istituto di Microbiologia, Facolta' di Medicina e Chirurgia, Bari-70124, Italy [P. P. D. F.]; Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232 [J. B., G. C.]; and the Kimmel Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 19107 [I. G., O. B. G., J. H. K.]

eps15, a substrate for the epidermal growth factor receptor and other receptor tyrosine kinases, possesses a discrete domain structure with protein-binding properties. It interacts with a number of cellular proteins through an evolutionarily conserved protein-binding domain, the eps15 homology domain, located in its NH₂-terminal region. In addition, a proline-rich region, located in the COOH-terminal portion of eps15, can bind to the Src homology 3 domain of the crk proto-oncogene product in vitro. Recently, coimmunoprecipitation between eps15 and AP-2, a major component of coated pits, was reported. Here, we characterize the molecular determinants of the eps15/AP-2 interaction. The AP-2 binding region of eps15 is localized in its COOH-terminal region and spans 80 amino acids. At least three molecular determinants, located at residues 650–660, 680–690, and 720–730, are involved in the binding. AP-2 binds to eps15 through its subunit (-adaptin); in particular, the COOH-terminal region of -adaptin, the so-called -ear, contains the eps15 binding region.

¹ This work was supported in part by grants from Consiglio Nazionale delle Ricerche and Associazione Italiana Ricerca sul Cancro (to P. P. D. F. and P. G. P.) and by NIH Grants GM-49217 (to J. H. K.) and CA-24071 (to G. C.). G. I. was supported by an Associazione Italiana Ricerca sul Cancro fellowship and European Community, project BIOMED-2. O. G. was supported by NIH Training Grant 5-T32-DK07705. J. B. was supported by a fellowship from the Spanish Ministerio de Education y Ciencia.

² To whom requests for reprints should be addressed, at the Kimmel Cancer Institute, Thomas Jefferson University, 233 South 10th Street, BLSB/915, Philadelphia, PA 19107. Phone: (215) 503-4624; Fax: (215) 923-1098; E-mail: j_keen@lac.jci.tju.edu.

Received 6/27/96. Accepted 11/18/96.

This article has been cited by other articles:

				S. Chi, H. Cao, J. Chen, and M. A. McNiven Eps15 Mediates Vesicle Trafficking from the trans-Golgi Network via an Interaction with the Clathrin Adaptor AP-1 Mol. Biol. Cell, August 1, 2008; 19(8): 3564 - 3575. [Abstract] [Full Text] [PDF]

				M. A. Brindley and W. Maury Equine Infectious Anemia Virus Entry Occurs through Clathrin-Mediated Endocytosis J. Virol., February 15, 2008; 82(4): 1628 - 1637. [Abstract] [Full Text] [PDF]

				I. Stavrou and T. J. O'Halloran The Monomeric Clathrin Assembly Protein, AP180, Regulates Contractile Vacuole Size in Dictyostelium discoideum Mol. Biol. Cell, December 1, 2006; 17(12): 5381 - 5389. [Abstract] [Full Text] [PDF]

				C. Clement, V. Tiwari, P. M. Scanlan, T. Valyi-Nagy, B. Y.J.T. Yue, and D. Shukla A novel role for phagocytosis-like uptake in herpes simplex virus entry J. Cell Biol., September 25, 2006; 174(7): 1009 - 1021. [Abstract] [Full Text] [PDF]

				M. A. McNiven and H. M. Thompson Vesicle formation at the plasma membrane and trans-Golgi network: the same but different. Science, September 15, 2006; 313(5793): 1591 - 1594. [Abstract] [Full Text] [PDF]

				T. Oshiro, S. Koyama, S. Sugiyama, A. Kondo, Y. Onodera, T. Asahara, H. Sabe, and A. Kikuchi Interaction of POB1, a Downstream Molecule of Small G Protein Ral, with PAG2, a Paxillin-binding Protein, Is Involved in Cell Migration J. Biol. Chem., October 4, 2002; 277(41): 38618 - 38626. [Abstract] [Full Text] [PDF]

				S. B. Sieczkarski and G. R. Whittaker Influenza Virus Can Enter and Infect Cells in the Absence of Clathrin-Mediated Endocytosis J. Virol., September 11, 2002; 76(20): 10455 - 10464. [Abstract] [Full Text] [PDF]

				E. Klapisz, I. Sorokina, S. Lemeer, M. Pijnenburg, A. J. Verkleij, and P. M. P. van Bergen en Henegouwen A Ubiquitin-interacting Motif (UIM) Is Essential for Eps15 and Eps15R Ubiquitination J. Biol. Chem., August 16, 2002; 277(34): 30746 - 30753. [Abstract] [Full Text] [PDF]

				K. R. Henry, K. D'Hondt, J. Chang, T. Newpher, K. Huang, R. T. Hudson, H. Riezman, and S. K. Lemmon Scd5p and Clathrin Function Are Important for Cortical Actin Organization, Endocytosis, and Localization of Sla2p in Yeast Mol. Biol. Cell, August 1, 2002; 13(8): 2607 - 2625. [Abstract] [Full Text] [PDF]

				E. S. Seto, H. J. Bellen, and T. E. Lloyd When cell biology meets development: endocytic regulation of signaling pathways Genes & Dev., June 1, 2002; 16(11): 1314 - 1336. [Full Text] [PDF]

				G. Baillat, S. Gaillard, F. Castets, and A. Monneron Interactions of Phocein with Nucleoside-Diphosphate Kinase, Eps15, and Dynamin I J. Biol. Chem., May 17, 2002; 277(21): 18961 - 18966. [Abstract] [Full Text] [PDF]

				V. Poupon, S. Polo, M. Vecchi, G. Martin, A. Dautry-Varsat, N. Cerf-Bensussan, P. P. Di Fiore, and A. Benmerah Differential Nucleocytoplasmic Trafficking between the Related Endocytic Proteins Eps15 and Eps15R J. Biol. Chem., March 8, 2002; 277(11): 8941 - 8948. [Abstract] [Full Text] [PDF]

				E. Santolini, C. Puri, A. E. Salcini, M. C. Gagliani, P. G. Pelicci, C. Tacchetti, and P. P. Di Fiore Numb Is an Endocytic Protein J. Cell Biol., December 11, 2000; 151(6): 1345 - 1352. [Abstract] [Full Text] [PDF]

				S. Confalonieri, A. E. Salcini, C. Puri, C. Tacchetti, and P. P. Di Fiore Tyrosine Phosphorylation of Eps15 Is Required for Ligand-Regulated, but Not Constitutive, Endocytosis J. Cell Biol., August 21, 2000; 150(4): 905 - 912. [Abstract] [Full Text] [PDF]

				R. Fernandez-Chacon, M. Achiriloaie, R. Janz, J. P. Albanesi, and T. C. Sudhof SCAMP1 Function in Endocytosis J. Biol. Chem., April 21, 2000; 275(17): 12752 - 12756. [Abstract] [Full Text] [PDF]

				J. Domin, I. Gaidarov, M. E. K. Smith, J. H. Keen, and M. D. Waterfield The Class II Phosphoinositide 3-Kinase PI3K-C2alpha Is Concentrated in the Trans-Golgi Network and Present in Clathrin-coated Vesicles J. Biol. Chem., April 14, 2000; 275(16): 11943 - 11950. [Abstract] [Full Text] [PDF]

				A. Benmerah, V. Poupon, N. Cerf-Bensussan, and A. Dautry-Varsat Mapping of Eps15 Domains Involved in Its Targeting to Clathrin-coated Pits J. Biol. Chem., February 4, 2000; 275(5): 3288 - 3295. [Abstract] [Full Text] [PDF]

				B Gagny, A Wiederkehr, P Dumoulin, B Winsor, H Riezman, and R Haguenauer-Tsapis A novel EH domain protein of Saccharomyces cerevisiae, Ede1p, involved in endocytosis J. Cell Sci., January 9, 2000; 113(18): 3309 - 3319. [Abstract] [PDF]

				A. Cadavid, A Ginzel, and J. Fischer The function of the Drosophila fat facets deubiquitinating enzyme in limiting photoreceptor cell number is intimately associated with endocytosis Development, January 4, 2000; 127(8): 1727 - 1736. [Abstract] [PDF]

				L. J. Page, P. J. Sowerby, W. W.Y. Lui, and M. S. Robinson {gamma}-Synergin: An Eh Domain-Containing Protein That Interacts with {gamma}-Adaptin J. Cell Biol., September 6, 1999; 146(5): 993 - 1004. [Abstract] [Full Text] [PDF]

				I. Gaidarov and J. H. Keen Phosphoinositide-Ap-2 Interactions Required for Targeting to Plasma Membrane Clathrin-Coated Pits J. Cell Biol., August 23, 1999; 146(4): 755 - 764. [Abstract] [Full Text] [PDF]

				E. Ungewickell Wrapping the package PNAS, August 3, 1999; 96(16): 8809 - 8810. [Full Text] [PDF]

				L. M. Traub, M. A. Downs, J. L. Westrich, and D. H. Fremont Crystal structure of the alpha appendage of AP-2 reveals a recruitment platform for clathrin-coat assembly PNAS, August 3, 1999; 96(16): 8907 - 8912. [Abstract] [Full Text] [PDF]

				V. Poupon, B. Begue, J. Gagnon, A. Dautry-Varsat, N. Cerf-Bensussan, and A. Benmerah Molecular Cloning and Characterization of MT-ACT48, a Novel Mitochondrial Acyl-CoA Thioesterase J. Biol. Chem., July 2, 1999; 274(27): 19188 - 19194. [Abstract] [Full Text] [PDF]

				M. Okamoto, S. Schoch, and T. C. Sudhof EHSH1/Intersectin, a Protein That Contains EH and SH3 Domains and Binds to Dynamin and SNAP-25. A PROTEIN CONNECTION BETWEEN EXOCYTOSIS AND ENDOCYTOSIS? J. Biol. Chem., June 25, 1999; 274(26): 18446 - 18454. [Abstract] [Full Text] [PDF]

				N. K. Hussain, M. Yamabhai, A. R. Ramjaun, A. M. Guy, D. Baranes, J. P. O'Bryan, C. J. Der, B. K. Kay, and P. S. McPherson Splice Variants of Intersectin Are Components of the Endocytic Machinery in Neurons and Nonneuronal Cells J. Biol. Chem., May 28, 1999; 274(22): 15671 - 15677. [Abstract] [Full Text] [PDF]

				H. Chen, V. I. Slepnev, P. P. Di Fiore, and P. De Camilli The Interaction of Epsin and Eps15 with the Clathrin Adaptor AP-2 Is Inhibited by Mitotic Phosphorylation and Enhanced by Stimulation-dependent Dephosphorylation in Nerve Terminals J. Biol. Chem., February 5, 1999; 274(6): 3257 - 3260. [Abstract] [Full Text] [PDF]

				M. R. Torrisi, L. V. Lotti, F. Belleudi, R. Gradini, A. E. Salcini, S. Confalonieri, P. G. Pelicci, and P. P. Di Fiore Eps15 Is Recruited to the Plasma Membrane upon Epidermal Growth Factor Receptor Activation and Localizes to Components of the Endocytic Pathway during Receptor Internalization Mol. Biol. Cell, February 1, 1999; 10(2): 417 - 434. [Abstract] [Full Text]

				A Benmerah, M Bayrou, N Cerf-Bensussan, and A Dautry-Varsat Inhibition of clathrin-coated pit assembly by an Eps15 mutant J. Cell Sci., January 5, 1999; 112(9): 1303 - 1311. [Abstract] [PDF]

				H Boleti, A Benmerah, D. Ojcius, N Cerf-Bensussan, and A Dautry-Varsat Chlamydia infection of epithelial cells expressing dynamin and Eps15 mutants: clathrin-independent entry into cells and dynamin-dependent productive growth J. Cell Sci., January 5, 1999; 112(10): 1487 - 1496. [Abstract] [PDF]

				T Sorkina, A Bild, F Tebar, and A Sorkin Clathrin, adaptors and eps15 in endosomes containing activated epidermal growth factor receptors J. Cell Sci., January 2, 1999; 112(3): 317 - 327. [Abstract] [PDF]

				J. Roos and R. B. Kelly Dap160, a Neural-specific Eps15 Homology and Multiple SH3 Domain-containing Protein That Interacts with Drosophila Dynamin J. Biol. Chem., July 24, 1998; 273(30): 19108 - 19119. [Abstract] [Full Text] [PDF]

				A. Benmerah, C. Lamaze, B. Begue, S. L. Schmid, A. Dautry-Varsat, and N. Cerf-Bensussan AP-2/Eps15 Interaction Is Required for Receptor-mediated Endocytosis J. Cell Biol., March 9, 1998; 140(5): 1055 - 1062. [Abstract] [Full Text] [PDF]

				L. Coda, A. E. Salcini, S. Confalonieri, G. Pelicci, T. Sorkina, A. Sorkin, P. G. Pelicci, and P. P. Di Fiore Eps15R Is a Tyrosine Kinase Substrate with Characteristics of a Docking Protein Possibly Involved in Coated Pits-mediated Internalization J. Biol. Chem., January 30, 1998; 273(5): 3003 - 3012. [Abstract] [Full Text] [PDF]

				P. Cupers, A. P. Jadhav, and T. Kirchhausen Assembly of Clathrin Coats Disrupts the Association between Eps15 and AP-2 Adaptors J. Biol. Chem., January 23, 1998; 273(4): 1847 - 1850. [Abstract] [Full Text] [PDF]

				P. Cupers, E. ter Haar, W. Boll, and T. Kirchhausen Parallel Dimers and Anti-parallel Tetramers Formed by Epidermal Growth Factor Receptor Pathway Substrate Clone 15 (EPS15) J. Biol. Chem., December 26, 1997; 272(52): 33430 - 33434. [Abstract] [Full Text] [PDF]

				A. Yamaguchi, T. Urano, T. Goi, and L. A. Feig An Eps Homology (EH) Domain Protein That Binds to the Ral-GTPase Target, RalBP1 J. Biol. Chem., December 12, 1997; 272(50): 31230 - 31234. [Abstract] [Full Text] [PDF]

				A. E. Salcini, S. Confalonieri, M. Doria, E. Santolini, E. Tassi, O. Minenkova, G. Cesareni, P. G. Pelicci, and P. P. Di Fiore Binding specificity and in vivo targets of the EH domain, a novel protein-protein interaction module Genes & Dev., September 1, 1997; 11(17): 2239 - 2249. [Abstract] [Full Text] [PDF]

				F. Tebar, S. Confalonieri, R. E. Carter, P. P. Di Fiore, and A. Sorkin Eps15 Is Constitutively Oligomerized Due to Homophilic Interaction of Its Coiled-coil Region J. Biol. Chem., June 13, 1997; 272(24): 15413 - 15418. [Abstract] [Full Text] [PDF]

				K. Kariya, S. Koyama, S. Nakashima, T. Oshiro, K. Morinaka, and A. Kikuchi Regulation of Complex Formation of POB1/Epsin/Adaptor Protein Complex 2 by Mitotic Phosphorylation J. Biol. Chem., June 9, 2000; 275(24): 18399 - 18406. [Abstract] [Full Text] [PDF]

				A. Fingerhut, K. von Figura, and S. Honing Binding of AP2 to Sorting Signals Is Modulated by AP2 Phosphorylation J. Biol. Chem., February 16, 2001; 276(8): 5476 - 5482. [Abstract] [Full Text] [PDF]

				V. I. Slepnev, G.-C. Ochoa, M. H. Butler, and P. De Camilli Tandem Arrangement of the Clathrin and AP-2 Binding Domains in Amphiphysin 1 and Disruption of Clathrin Coat Function by Amphiphysin Fragments Comprising These Sites J. Biol. Chem., June 2, 2000; 275(23): 17583 - 17589. [Abstract] [Full Text] [PDF]