title: Rac1 recruits the adapter protein CMS/CD2AP to cell-cell contacts.
abstract: Rac1 is a member of the Rho family of small GTPases, which regulate cell adhesion and migration through their control of the actin cytoskeleton. Rho-GTPases are structurally very similar, with the exception of a hypervariable domain in the C terminus. Using peptide-based pulldown assays in combination with mass spectrometry, we previously showed that the hypervariable domain in Rac1 mediates specific protein-protein interactions. Most recently, we found that the Rac1 C terminus associates to the ubiquitously expressed adapter protein CMS/CD2AP. CD2AP is critical for the formation and maintenance of a specialized cell-cell contact between kidney podocyte foot processes, the slit diaphragm. Here, CD2AP links the cell adhesion protein nephrin to the actin cytoskeleton. In addition, CMS/CD2AP binds actin-regulating proteins, such as CAPZ and cortactin, and has been implicated in the internalization of growth factor receptors. We found that CD2AP specifically interacts with the C-terminal domain of Rac1 but not with that of other Rho family members. Efficient interaction between Rac1 and CD2AP requires both the proline-rich domain and the poly-basic region in the Rac1 C terminus, and at least two of the three N-terminal SH3 domains of CD2AP. CD2AP co-localizes with Rac1 to membrane ruffles, and small interfering RNA-based experiments showed that CD2AP links Rac1 to CAPZ and cortactin. Finally, expression of constitutive active Rac1 recruits CD2AP to cell-cell contacts in epithelial cells, where we found CD2AP to participate in the control of the epithelial barrier function. These data identify CD2AP as a novel Rac1-associated adapter protein that participates in the regulation of epithelial cell-cell contact.
authors.affiliation: Department of Molecular Cell Biology, Sanquin Research and Landsteiner Laboratory, University of Amsterdam, 1066 CX, Amsterdam.
authors.forename: Trynette J
authors.initials: TJ
authors.lastname: van Duijn
authors.forename: Eloise C
authors.initials: EC
authors.lastname: Anthony
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Hensbergen
authors.forename: André M
authors.initials: AM
authors.lastname: Deelder
authors.forename: Peter L
authors.initials: PL
authors.lastname: Hordijk
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D051347:CapZ Actin Capping Protein / Q000235:genetics / Q000378:metabolism; D002448:Cell Adhesion; D002450:Cell Communication; D002460:Cell Line; D045744:Cell Line, Tumor; D051356:Cortactin / Q000235:genetics / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D019169:Jurkat Cells; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding; D025941:Protein Interaction Mapping; D034622:RNA Interference; D014162:Transfection; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D018909:src Homology Domains
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C495842:CAPZA1 protein, human; C114545:CD2-associated protein; D051347:CapZ Actin Capping Protein; D051356:Cortactin; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; C113267:nephrin; D049452:Green Fluorescent Proteins; D020830:rac1 GTP-Binding Protein
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