|
|
|
|
|
||
|
|||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | |||||
Research Article |
Department of Cell Biology, University of Virginia, Charlottesville VA
22908
* Present address: Ri-chiroh Manabe, Japan Science and Technology Corporation,
Sekiguchi Biomatrix Signaling Project, c/o Aichi Medical University, 21
Karimata-Yasago, Nagakute, Aichi 480-1195, Japan
These authors contributed equally to this work
Author for correspondence (e-mail:
horwitz{at}virginia.edu
)
Accepted 23 January 2002
GIT1 is a multidomain protein that is thought to function as an integrator of signaling pathways controlling vesicle trafficking, adhesion and cytoskeletal organization. It regulates ARF GTPases and has binding domains for paxillin and PIX, which is a PAK-binding protein and an exchange factor for Rac. We show that GIT1 cycles between at least three distinct subcellular compartments, including adhesion-like structures, the leading edge and cytoplasmic complexes. The cytoplasmic structures, which also contain paxillin, PAK and PIX, do not detectably co-localize with endosomal Golgi or membrane markers, suggesting that they represent a novel supramolecular complex. The GIT1 cytoplasmic complexes are motile and tended to move toward the cell periphery where they joined existing adhesions. In retracting regions of the cells, the GIT1 complexes moved away from the disassembling adhesions toward the cell body. Using deletion mutants, we have identified domains that target GIT1 to each of the compartments. Localization to adhesions and the leading edge requires the paxillin-binding domain, which comprises the C-terminal 140 residues (cGIT1), whereas targeting to the cytoplasmic complexes requires the central region that contains ankyrin repeats and the PIX-binding domain. Expression of GIT1 or cGIT, but not nGIT1 in which the paxillin-binding domain is deleted, increases the rate of migration and the size and number of protrusions. The latter are inhibited when GIT1 is co-expressed with a kinase-dead PAK, suggesting that the GIT1 interaction with PAK is required for enhanced migration and protrusive activity. Furthermore, GIT1 targets constitutively activated PAK to adhesions and the leading edge via its interaction with paxillin. Since expression of cGIT targets endogenous GIT1 to the leading edge, it appears that the leading edge is the location of GIT1 responsible for these activities. Thus, GIT1 is a component of a motile, multimolecular complex that traffics a set of signaling components to specific locations in the cell where they regulate localized activities.
Key words: Cell adhesion, GFP, GIT1, p21 activated kinase (PAK), Rac, Paxillin
This article has been cited by other articles:
|
|
 |
|
  M. Vicente-Manzanares, C. K. Choi, and A. R. Horwitz Integrins in cell migration - the actin connection J. Cell Sci., January 15, 2009; 122(2): 199 - 206. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Nola, M. Sebbagh, S. Marchetto, N. Osmani, C. Nourry, S. Audebert, C. Navarro, R. Rachel, M. Montcouquiol, N. Sans, et al. Scrib regulates PAK activity during the cell migration process Hum. Mol. Genet., November 15, 2008; 17(22): 3552 - 3565. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. E. Cavet, J. Pang, G. Yin, and B. C. Berk An epidermal growth factor (EGF) -dependent interaction between GIT1 and sorting nexin 6 promotes degradation of the EGF receptor FASEB J, October 1, 2008; 22(10): 3607 - 3616. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. J. Coniglio, S. Zavarella, and M. H. Symons Pak1 and Pak2 Mediate Tumor Cell Invasion through Distinct Signaling Mechanisms Mol. Cell. Biol., June 15, 2008; 28(12): 4162 - 4172. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. L. Ha, S. Bharti, H. Inoue, W. C. Vass, F. Campa, Z. Nie, A. de Gramont, Y. Ward, and P. A. Randazzo ASAP3 Is a Focal Adhesion-associated Arf GAP That Functions in Cell Migration and Invasion J. Biol. Chem., May 30, 2008; 283(22): 14915 - 14926. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Chahdi and A. Sorokin Protein Kinase A-Dependent Phosphorylation Modulates {beta}1Pix Guanine Nucleotide Exchange Factor Activity through 14-3-3{beta} Binding Mol. Cell. Biol., March 1, 2008; 28(5): 1679 - 1687. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Totaro, S. Paris, C. Asperti, and I. de Curtis Identification of an Intramolecular Interaction Important for the Regulation of GIT1 Functions Mol. Biol. Cell, December 1, 2007; 18(12): 5124 - 5138. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. W. Mayhew, E. D. Jeffery, N. E. Sherman, K. Nelson, J. M. Polefrone, S. J. Pratt, J. Shabanowitz, J. T. Parsons, J. W. Fox, D. F. Hunt, et al. Identification of phosphorylation sites in betaPIX and PAK1 J. Cell Sci., November 15, 2007; 120(22): 3911 - 3918. [Full Text] [PDF]
|
|
|
|
|
|
N. P. Jones and M. Katan Role of Phospholipase C{gamma}1 in Cell Spreading Requires Association with a {beta}-Pix/GIT1-Containing Complex, Leading to Activation of Cdc42 and Rac1 Mol. Cell. Biol., August 15, 2007; 27(16): 5790 - 5805. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. T. Gerthoffer Mechanisms of Vascular Smooth Muscle Cell Migration Circ. Res., March 16, 2007; 100(5): 607 - 621. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-C. Shen, M. Unoki, D. Ythier, A. Duperray, L. Varticovski, K. Kumamoto, R. Pedeux, and C. C. Harris Inhibitor of Growth 4 Suppresses Cell Spreading and Cell Migration by Interacting with a Novel Binding Partner, Liprin {alpha}1 Cancer Res., March 15, 2007; 67(6): 2552 - 2558. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. J. Webb, H. Zhang, D. Majumdar, and A. F. Horwitz {alpha}5 Integrin Signaling Regulates the Formation of Spines and Synapses in Hippocampal Neurons J. Biol. Chem., March 9, 2007; 282(10): 6929 - 6935. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Hajdo-Milasinovic, S. I. J. Ellenbroek, S. van Es, B. van der Vaart, and J. G. Collard Rac1 and Rac3 have opposing functions in cell adhesion and differentiation of neuronal cells J. Cell Sci., February 15, 2007; 120(4): 555 - 566. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Chang, C. A. Lemmon, D. Park, and L. H. Romer FAK Potentiates Rac1 Activation and Localization to Matrix Adhesion Sites: A Role for betaPIX Mol. Biol. Cell, January 1, 2007; 18(1): 253 - 264. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. J. Webb, M. W. Mayhew, M. Kovalenko, M. J. Schroeder, E. D. Jeffery, L. Whitmore, J. Shabanowitz, D. F. Hunt, and A. F. Horwitz Identification of phosphorylation sites in GIT1 J. Cell Sci., July 15, 2006; 119(14): 2847 - 2850. [Full Text] [PDF]
|
|
|
|
|
|
S. Frese, W.-D. Schubert, A. C. Findeis, T. Marquardt, Y. S. Roske, T. E. B. Stradal, and D. W. Heinz The Phosphotyrosine Peptide Binding Specificity of Nck1 and Nck2 Src Homology 2 Domains J. Biol. Chem., June 30, 2006; 281(26): 18236 - 18245. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Nayal, D. J. Webb, C. M. Brown, E. M. Schaefer, M. Vicente-Manzanares, and A. R. Horwitz Paxillin phosphorylation at Ser273 localizes a GIT1-PIX-PAK complex and regulates adhesion and protrusion dynamics J. Cell Biol., May 22, 2006; 173(4): 587 - 589. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. J. Hoefen and B. C. Berk The multifunctional GIT family of proteins. J. Cell Sci., April 15, 2006; 119(Pt 8): 1469 - 1475. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. E. Tcheperegine, X.-D. Gao, and E. Bi Regulation of Cell Polarity by Interactions of Msb3 and Msb4 with Cdc42 and Polarisome Components Mol. Cell. Biol., October 1, 2005; 25(19): 8567 - 8580. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Brown, L. A. Cary, J. S. Jamieson, J. A. Cooper, and C. E. Turner Src and FAK Kinases Cooperate to Phosphorylate Paxillin Kinase Linker, Stimulate Its Focal Adhesion Localization, and Regulate Cell Spreading and Protrusiveness Mol. Biol. Cell, September 1, 2005; 16(9): 4316 - 4328. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Yin, Q. Zheng, C. Yan, and B. C. Berk GIT1 Is a Scaffold for ERK1/2 Activation in Focal Adhesions J. Biol. Chem., July 29, 2005; 280(30): 27705 - 27712. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. T. Barfod, A. L. Moore, R. F. Melnick, and S. D. Lidofsky Src Regulates Distinct Pathways for Cell Volume Control through Vav and Phospholipase C{gamma} J. Biol. Chem., July 8, 2005; 280(27): 25548 - 25557. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. P. Jones, J. Peak, S. Brader, S. A. Eccles, and M. Katan PLC{gamma}1 is essential for early events in integrin signalling required for cell motility J. Cell Sci., June 15, 2005; 118(12): 2695 - 2706. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Cau and A. Hall Cdc42 controls the polarity of the actin and microtubule cytoskeletons through two distinct signal transduction pathways J. Cell Sci., June 15, 2005; 118(12): 2579 - 2587. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Zhang, D. J. Webb, H. Asmussen, S. Niu, and A. F. Horwitz A GIT1/PIX/Rac/PAK Signaling Module Regulates Spine Morphogenesis and Synapse Formation through MLC J. Neurosci., March 30, 2005; 25(13): 3379 - 3388. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Rosenberger, A. Gal, and K. Kutsche {alpha}PIX Associates with Calpain 4, the Small Subunit of Calpain, and Has a Dual Role in Integrin-mediated Cell Spreading J. Biol. Chem., February 25, 2005; 280(8): 6879 - 6889. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. M. Hinsby, J. V. Olsen, and M. Mann Tyrosine Phosphoproteomics of Fibroblast Growth Factor Signaling: A ROLE FOR INSULIN RECEPTOR SUBSTRATE-4 J. Biol. Chem., November 5, 2004; 279(45): 46438 - 46447. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. C. Brown and C. E. Turner Paxillin: Adapting to Change Physiol Rev, October 1, 2004; 84(4): 1315 - 1339. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. R. Stofega, L. C. Sanders, E. M. Gardiner, and G. M. Bokoch Constitutive p21-activated Kinase (PAK) Activation in Breast Cancer Cells as a Result of Mislocalization of PAK to Focal Adhesions Mol. Biol. Cell, June 1, 2004; 15(6): 2965 - 2977. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T.-H. Loo, Y.-W. Ng, L. Lim, and E. Manser GIT1 Activates p21-Activated Kinase through a Mechanism Independent of p21 Binding Mol. Cell. Biol., May 1, 2004; 24(9): 3849 - 3859. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E.-Y. Shin, K.-N. Woo, C.-S. Lee, S.-H. Koo, Y. G. Kim, W.-J. Kim, C.-D. Bae, S.-I. Chang, and E.-G. Kim Basic Fibroblast Growth Factor Stimulates Activation of Rac1 through a p85 {beta}PIX Phosphorylation-dependent Pathway J. Biol. Chem., January 16, 2004; 279(3): 1994 - 2004. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Lamorte, S. Rodrigues, V. Sangwan, C. E. Turner, and M. Park Crk Associates with a Multimolecular Paxillin/GIT2/{beta}-PIX Complex and Promotes Rac-dependent Relocalization of Paxillin to Focal Contacts Mol. Biol. Cell, July 1, 2003; 14(7): 2818 - 2831. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Zhang, D. J. Webb, H. Asmussen, and A. F. Horwitz Synapse formation is regulated by the signaling adaptor GIT1 J. Cell Biol., April 14, 2003; 161(1): 131 - 142. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Ko, S. Kim, J. G. Valtschanoff, H. Shin, J.-R. Lee, M. Sheng, R. T. Premont, R. J. Weinberg, and E. Kim Interaction between Liprin-alpha and GIT1 Is Required for AMPA Receptor Targeting J. Neurosci., March 1, 2003; 23(5): 1667 - 1677. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Kim, J. Ko, H. Shin, J.-R. Lee, C. Lim, J.-H. Han, W. D. Altrock, C. C. Garner, E. D. Gundelfinger, R. T. Premont, et al. The GIT Family of Proteins Forms Multimers and Associates with the Presynaptic Cytomatrix Protein Piccolo J. Biol. Chem., February 14, 2003; 278(8): 6291 - 6300. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Mellman and A. Ridley Regulation and Functional Insights in Cellular Polarity Madrid, Spain June 3-5, 2002 J. Cell Biol., July 8, 2002; 158(1): 12 - 16. [Full Text] [PDF]
|
|
