Rac1 recruits the adapter protein CMS/CD2AP to cell-cell contacts.
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.
van Duijn
Trynette J
TJ
Department of Molecular Cell Biology, Sanquin Research and Landsteiner Laboratory, University of Amsterdam, 1066 CX, Amsterdam.
Anthony
Eloise C
EC
Hensbergen
Paul J
PJ
Deelder
André M
AM
Hordijk
Peter L
PL
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
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
J Cell Biol. 2001 Jan 8;152(1):111-26
11149925
Nat Rev Immunol. 2009 Sep;9(9):630-44
19696767
EMBO J. 2007 Jan 24;26(2):336-45
17245428
Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7896-900
1881923
Nat Rev Mol Cell Biol. 2005 Feb;6(2):167-80
15688002
Biol Cell. 2007 Jan;99(1):1-12
17155934
Proc Natl Acad Sci U S A. 1999 May 25;96(11):6211-6
10339567
J Cell Sci. 2007 Feb 15;120(Pt 4):555-66
17244648
J Cell Sci. 2007 Jan 1;120(Pt 1):86-100
17164293
Am J Physiol Lung Cell Mol Physiol. 2001 Dec;281(6):L1500-11
11704547
J Biol Chem. 2004 Aug 20;279(34):35127-32
15166221
Am J Physiol Renal Physiol. 2002 Oct;283(4):F734-43
12217865
J Biol Chem. 2001 Feb 16;276(7):4957-63
11067845
Cell. 1998 Sep 4;94(5):667-77
9741631
Lancet. 2003 Nov 22;362(9397):1746-8
14643126
Nephrology (Carlton). 2006 Aug;11(4):274-81
16889564
FEBS Lett. 2003 Dec 18;555(3):556-60
14675773
J Biol Chem. 2006 Dec 15;281(50):38845-53
17020880
Am J Physiol Renal Physiol. 2000 Oct;279(4):F785-92
10997929
Biochim Biophys Acta. 2009 Apr;1788(4):790-6
19159611
J Cell Biol. 2004 Oct 11;167(1):111-22
15479739
Curr Biol. 2002 Mar 5;12(5):379-82
11882288
J Biol Chem. 2003 Oct 3;278(40):39166-75
12874273
Am J Physiol Renal Physiol. 2001 Oct;281(4):F769-77
11553524
Biochim Biophys Acta. 2009 Apr;1788(4):797-812
19366594
Mol Biol Cell. 2004 Mar;15(3):1065-76
14699063
J Cell Sci. 2001 Aug;114(Pt 15):2713-22
11683406
Exp Nephrol. 2001;9(6):355-9
11701993
Cell. 1992 Aug 7;70(3):401-10
1643658
J Biol Chem. 2003 Jun 13;278(24):21805-13
12672817
Am J Pathol. 2001 Dec;159(6):2303-8
11733379
J Biol Chem. 2008 Nov 14;283(46):31830-9
18809679
Science. 1997 Nov 21;278(5342):1464-6
9367959
J Cell Biol. 2006 Feb 27;172(5):759-69
16492808
Am J Physiol Renal Physiol. 2002 Apr;282(4):F585-91
11880318
J Cell Biol. 2008 Mar 24;180(6):1191-204
18362180
Proc Natl Acad Sci U S A. 2000 May 9;97(10):5225-30
10805781
Cancer Res. 2001 Jan 15;61(2):474-7
11212234
J Biol Chem. 2006 Jul 14;281(28):19196-203
16707503
J Biol Chem. 2003 Jun 20;278(25):22396-403
12690097
Eur J Cell Biol. 2008 Oct;87(10):779-92
18635287
J Biol Chem. 1997 Sep 12;272(37):23371-5
9287351
J Biol Chem. 2004 Aug 20;279(34):35840-8
15143066
Blood. 2001 Feb 1;97(3):799-804
11157500
J Biol Chem. 2003 Oct 10;278(41):39735-46
12874286
Nephrol Dial Transplant. 2009 Jun;24(6):1858-64
19131354
Science. 1999 Oct 8;286(5438):312-5
10514378
Nat Cell Biol. 2002 Mar;4(3):232-9
11862216
Nephron Exp Nephrol. 2010;114(3):e93-e106
19955829
Biochem Biophys Res Commun. 2005 Jul 1;332(2):426-32
15910750