title: The residence time of focal adhesion kinase (FAK) and paxillin at focal adhesions in renal epithelial cells is determined by adhesion size, strength and life cycle status.
abstract: Focal adhesions (FAs) are specialized membrane-associated multi-protein complexes that link the cell to the extracellular matrix and enable cell proliferation, survival and motility. Despite the extensive description of the molecular composition of FAs, the complex regulation of FA dynamics is unclear. We have used photobleaching assays of whole cells to determine the protein dynamics in every single focal adhesion. We identified that the focal adhesion proteins FAK and paxillin exist in two different states: a diffuse cytoplasmic pool and a transiently immobile FA-bound fraction with variable residence times. Interestingly, the average residence time of both proteins increased with focal adhesion size. Moreover, increasing integrin clustering by modulating surface collagen density increased residence time of FAK but not paxillin. Finally, this approach was applied to measure FAK and paxillin dynamics using nocodazole treatment followed by washout. This revealed an opposite residence time of FAK and paxillin in maturing and disassembling FAs, which depends on the ventral and peripheral cellular position of the FAs.
authors: Division of Toxicology, Leiden/Amsterdam Center for Drug Research, Leiden University, Leiden, Gorlaeus Laboratoria, PO Box 9502, 2300 RA Leiden,  The Netherlands.
authors: Sylvia E
authors: SE
authors: Le Dévédec
authors: Bart
authors: B
authors: Geverts
authors: Hans
authors: H
authors: de Bont
authors: Kuan
authors: K
authors: Yan
authors: Fons J
authors: FJ
authors: Verbeek
authors: Adriaan B
authors: AB
authors: Houtsmuller
authors: Bob
authors: B
authors: van de Water
mesh_terms: D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D003198:Computer Simulation; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004058:Diffusion; D004847:Epithelial Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology*; D036681:Fluorescence Recovery After Photobleaching; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism*; D022001:Focal Adhesions / Q000187:drug effects / Q000201:enzymology*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007700:Kinetics; D018374:LLC-PK1 Cells; D008024:Ligands; D008954:Models, Biological; D009010:Monte Carlo Method; D015739:Nocodazole / Q000494:pharmacology; D051419:Paxillin / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013552:Swine; D013997:Time Factors
chemical_list: D008024:Ligands; D051419:Paxillin; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D051416:Focal Adhesion Protein-Tyrosine Kinases; D015739:Nocodazole