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Research Article |
1 Department of Pediatrics and Pharmacology, University of Wisconsin, 1300
University Avenue, University of Wisconsin Medical School, Madison, WI 53706,
USA
2 Institute of Molecular Biology, Austrian Academy of Sciences, Salzburg A-5020,
Austria
3 Department of Cell and Molecular Physiology, University of North Carolina,
Chapel Hill, NC 27599, USA
* Author for correspondence (e-mail: huttenlocher{at}facstaff.wisc.edu)
Accepted 12 June 2002
Cell migration requires the regulated and dynamic turnover of adhesive
complexes. We have previously demonstrated that the calcium-dependent
protease, calpain, regulates the organization of adhesive complexes and cell
detachment during cell migration. Evidence is now provided that inhibiting
calpain through over-expression of the endogenous inhibitor of calpain,
calpastatin, and pharmacological inhibitors results in an inhibition of
adhesive complex disassembly with stabilization of GFP-vinculin and
GFP/RFP-zyxin at the cell periphery. Calpain was also required for the
microtubule-mediated turnover of adhesive complex sites after nocodazole
wash-out, suggesting that calpain may mediate focal complex disassembly
downstream of microtubules. Using dual imaging of RFP-zyxin and
GFP—
-actinin, we observed a temporal and spatial relationship
between -actinin localization to focal contacts and the subsequent
disassembly or translocation of RFP-zyxin containing focal complexes in areas
of cell retraction. Calpain inhibition disrupted -actinin localization
to zyxin-containing focal contacts and focal complex disassembly or
translocation to the cell center. In addition, disrupting -actinin
localization to focal complexes through expression of the -actinin rod
domain, but not the head domain, resulted in inhibition of focal adhesion
disassembly similar to calpain inhibition. Our studies suggest a novel
mechanism of action whereby calpain may modulate -actinin localization
into focal complexes and their subsequent disassembly or translocation.
Key words: Migration, Calpain, Cytoskeleton, Focal adhesion, -actinin
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