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Biophys J. 2008 Apr 1;94(7):2819-31. Epub 2007 Nov 9.

Paxillin dynamics measured during adhesion assembly and disassembly by correlation spectroscopy.

Author information

1
Laboratory for Fluorescence Dynamics, Department of Biomedical Engineering, University of California, Irvine, California, USA.

Erratum in

  • Biophys J. 2008 Jun;94(11):4577.

Abstract

Paxillin is an adaptor molecule involved in the assembly of focal adhesions. Using different fluorescence fluctuation approaches, we established that paxillin-EGFP is dynamic on many timescales within the cell, ranging from milliseconds to seconds. In the cytoplasmic regions, far from adhesions, paxillin is uniformly distributed and freely diffusing as a monomer, as determined by single-point fluctuation correlation spectroscopy and photon-counting histogram analysis. Near adhesions, paxillin dynamics are reduced drastically, presumably due to binding to protein partners within the adhesions. The photon-counting histogram analysis of the fluctuation amplitudes reveals that this binding equilibrium in new or assembling adhesions is due to paxillin monomers binding to quasi-immobile structures, whereas in disassembling adhesions or regions of adhesions, the equilibrium is due to exchange of large aggregates. Scanning fluctuation correlation spectroscopy and raster-scan image correlation spectroscopy analysis of laser confocal images show that the environments within adhesions are heterogeneous. Relatively large adhesions appear to slide transversally due to a treadmilling mechanism through the addition of monomeric paxillin at one side and removal of relatively large aggregates of proteins from the retracting edge. Total internal reflection microscopy performed with a fast acquisition EM-CCD camera completes the overall dynamic picture and adds details of the heterogeneous dynamics across single adhesions and simultaneous bursts of activity at many adhesions across the cell.

PMID:
17993500
PMCID:
PMC2267137
DOI:
10.1529/biophysj.107.104984
[Indexed for MEDLINE]
Free PMC Article

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