spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search    

The fully linked HTML version of this article has now been published.
JCS ePress online publication date 23 Jan 2003
doi: 10.1242/jcs.00284

This Article
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
jcs.00284v1
116/6/975    most recent
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kirchner, J.
Right arrow Articles by Geiger, B.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kirchner, J.
Right arrow Articles by Geiger, B.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Research Article

Live-cell monitoring of tyrosine phosphorylation in focal adhesions following microtubule disruption


Jochen Kirchner, Zvi Kam, Gila Tzur, Alexander D. Bershadsky, and Benjamin Geiger*
* Author for correspondence (e-mail: benny.geiger{at}weizmann.ac.il)

Tyrosine phosphorylation of focal adhesion components is involved in the regulation of focal adhesion formation and turnover, yet the underlying molecular mechanisms are still poorly defined. In the present study, we have used quantitative fluorescence microscopy to investigate the dynamic relationships between the incorporation of new components into growing focal adhesions and tyrosine phosphorylation of these sites. For this purpose, a new approach for monitoring phosphotyrosine levels in live cells was developed, based on a 'phosphotyrosine reporter' consisting of yellow fluorescent protein fused to two consecutive phosphotyrosine-binding Src-homology 2 (SH2)-domains derived from pp60c-Src. This YFP-dSH2 localized to cell-matrix adhesions and its intensity was linearly correlated with that of an anti-phosphotyrosine antibody labeling. The differential increase in vinculin and phosphotyrosine levels was examined in live cells by two-color time-lapse movies of CFP-vinculin and YFP-dSH2. In this study, focal adhesion growth was triggered by microtubule disruption, which was previously shown to stimulate focal adhesion development by inducing cellular contraction. We show here that, 2 minutes after addition of the microtubule-disrupting drug nocodazole, the local densities of the focal adhesion-associated proteins vinculin, paxillin and focal adhesion kinase (FAK) are significantly elevated and the focal adhesion area is increased, whereas elevation in tyrosine phosphorylation inside the growing adhesions occurs only a few minutes later. Phosphotyrosine and FAK density reach their maximum levels after 10 minutes of treatment, whereas vinculin and paxillin levels as well as focal adhesion size continue to grow, reaching a plateau at about 30 minutes. Our findings suggest that protein recruitment and growth of focal adhesions are an immediate and direct result of increased contractility induced by microtubule disruption, whereas tyrosine phosphorylation is activated later.
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?


This article has been cited by other articles:


Home page
 J. Neurosci.Home page
S. Woo, D. J. Rowan, and T. M. Gomez
Retinotopic Mapping Requires Focal Adhesion Kinase-Mediated Regulation of Growth Cone Adhesion
J. Neurosci., November 4, 2009; 29(44): 13981 - 13991.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
Q. Wang, D. Rajshankar, D. R. Branch, K. A. Siminovitch, M. T. H. Abreu, G. P. Downey, and C. A. McCulloch
Protein-tyrosine Phosphatase-{alpha} and Src Functionally Link Focal Adhesions to the Endoplasmic Reticulum to Mediate Interleukin-1-induced Ca2+ Signaling
J. Biol. Chem., July 31, 2009; 284(31): 20763 - 20772.
[Abstract] [Full Text] [PDF]

Home page
 J. Cell Sci.Home page
F. Stanchi, C. Grashoff, C. F. Nguemeni Yonga, D. Grall, R. Fassler, and E. Van Obberghen-Schilling
Molecular dissection of the ILK-PINCH-parvin triad reveals a fundamental role for the ILK kinase domain in the late stages of focal-adhesion maturation
J. Cell Sci., June 1, 2009; 122(11): 1800 - 1811.
[Abstract] [Full Text] [PDF]

Home page
 JCBHome page
K. T. Chan, C. L. Cortesio, and A. Huttenlocher
FAK alters invadopodia and focal adhesion composition and dynamics to regulate breast cancer invasion
J. Cell Biol., April 20, 2009; 185(2): 357 - 370.
[Abstract] [Full Text] [PDF]

Home page
 JCBHome page
Y. Lim, S.-T. Lim, A. Tomar, M. Gardel, J. A. Bernard-Trifilo, X. L. Chen, S. A. Uryu, R. Canete-Soler, J. Zhai, H. Lin, et al.
PyK2 and FAK connections to p190Rho guanine nucleotide exchange factor regulate RhoA activity, focal adhesion formation, and cell motility
J. Cell Biol., January 10, 2008; 180(1): 187 - 203.
[Abstract] [Full Text] [PDF]

Home page
 JCBHome page
J. D. Humphries, P. Wang, C. Streuli, B. Geiger, M. J. Humphries, and C. Ballestrem
Vinculin controls focal adhesion formation by direct interactions with talin and actin
J. Cell Biol., December 3, 2007; 179(5): 1043 - 1057.
[Abstract] [Full Text] [PDF]

Home page
 Mol. Biol. CellHome page
W.-h. Guo and Y.-l. Wang
Retrograde Fluxes of Focal Adhesion Proteins in Response to Cell Migration and Mechanical Signals
Mol. Biol. Cell, November 1, 2007; 18(11): 4519 - 4527.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
T. Schmitter, S. Pils, S. Weibel, F. Agerer, L. Peterson, A. Buntru, K. Kopp, and C. R. Hauck
Opa Proteins of Pathogenic Neisseriae Initiate Src Kinase-Dependent or Lipid Raft-Mediated Uptake via Distinct Human Carcinoembryonic Antigen-Related Cell Adhesion Molecule Isoforms
Infect. Immun., August 1, 2007; 75(8): 4116 - 4126.
[Abstract] [Full Text] [PDF]

Home page
 J. Cell Sci.Home page
A. D.-A. Tran, T. P. Marmo, A. A. Salam, S. Che, E. Finkelstein, R. Kabarriti, H. S. Xenias, R. Mazitschek, C. Hubbert, Y. Kawaguchi, et al.
HDAC6 deacetylation of tubulin modulates dynamics of cellular adhesions
J. Cell Sci., April 15, 2007; 120(8): 1469 - 1479.
[Abstract] [Full Text] [PDF]

Home page
 J. Pharmacol. Exp. Ther.Home page
R. G. Deschesnes, A. Patenaude, J. L. C. Rousseau, J. S. Fortin, C. Ricard, M.-F. Cote, J. Huot, R. C.-Gaudreault, and E. Petitclerc
Microtubule-Destabilizing Agents Induce Focal Adhesion Structure Disorganization and Anoikis in Cancer Cells
J. Pharmacol. Exp. Ther., February 1, 2007; 320(2): 853 - 864.
[Abstract] [Full Text] [PDF]

Home page
 J. Cell Sci.Home page
C. Ballestrem, N. Erez, J. Kirchner, Z. Kam, A. Bershadsky, and B. Geiger
Molecular mapping of tyrosine-phosphorylated proteins in focal adhesions using fluorescence resonance energy transfer
J. Cell Sci., March 1, 2006; 119(5): 866 - 875.
[Abstract] [Full Text] [PDF]

Home page
 J. Neurosci.Home page
E. Robles, S. Woo, and T. M. Gomez
Src-Dependent Tyrosine Phosphorylation at the Tips of Growth Cone Filopodia Promotes Extension
J. Neurosci., August 17, 2005; 25(33): 7669 - 7681.
[Abstract] [Full Text] [PDF]

Home page
 JCBHome page
A. Kodama, T. Lechler, and E. Fuchs
Coordinating cytoskeletal tracks to polarize cellular movements
J. Cell Biol., October 25, 2004; 167(2): 203 - 207.
[Abstract] [Full Text] [PDF]

Home page
 J. Cell Sci.Home page
F. Watt
Best Paper Award 2003
J. Cell Sci., April 15, 2004; 117(10): 1871 - 1871.
[Full Text] [PDF]


© The Company of Biologists Ltd 2003