Pak (p21-activated kinase) serine/threonine kinases have been shown to mediate directional sensing of chemokine gradients. We hypothesized that Pak may also mediate chemokine-induced shape changes, to facilitate leucocyte chemotaxis through restrictive barriers, such as the extracellular matrix. A potent inhibitor, Paki, was characterized and used to probe the role of Pak-family kinases in SDF-1α (stromal-cell derived factor-1α/CXCL12)-induced chemotaxis in a T cell model. Paki potently inhibited SDF-1α-induced Pak activation by a bivalent mechanism, as indicated by its complete inactivation upon point mutation of two binding sites, but partial inactivation upon mutation of either site alone. Importantly, Paki was not toxic to cells over the time frame of our experiments, since it did not substantially affect cell surface expression of CXCR4 (CXC chemokine receptor 4) or integrins, cell cycle progression, or a number of ligand-induced responses. Paki produced dose-dependent inhibition of SDF-1α-induced migration through rigid filters bearing small pores; but unexpectedly, did not substantially affect the magnitude or kinetics of chemotaxis through filters bearing larger pores. SDF-1α-induced Pak activation was partly dependent on PIX (Pak-interactive exchange factor); correspondingly, an allele of β-PIX that cannot bind Pak inhibited SDF-1α-induced chemotaxis through small, but not large pores. By contrast, other key players in chemotaxis: Gi, PI3K (phosphoinositide 3-kinase), and the Rho-family G-proteins, Rac and Cdc42 (cell division cycle 42), were required for SDF-1α-induced migration regardless of the barrier pore-size. These studies have revealed a distinct branch of the SDF-1α signalling pathway, in which the Rac/Cdc42 effector, Pak, and its partner, PIX, specifically regulate the cellular events required for chemokine-induced migration through restrictive barriers.
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Research Article|
June 14 2006
A Pak- and Pix-dependent branch of the SDF-1α signalling pathway mediates T cell chemotaxis across restrictive barriers
Natalia Volinsky;
Natalia Volinsky
1The Rappaport Family Institute for Research in the Medical Sciences, Bruce Rappaport Faculty of Medicine, Technion – Israel Institute of Technology, P.O. Box 9649, Bat Galim, Haifa 31096, Israel
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Anna Gantman;
Anna Gantman
1The Rappaport Family Institute for Research in the Medical Sciences, Bruce Rappaport Faculty of Medicine, Technion – Israel Institute of Technology, P.O. Box 9649, Bat Galim, Haifa 31096, Israel
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Deborah Yablonski
Deborah Yablonski
1
1The Rappaport Family Institute for Research in the Medical Sciences, Bruce Rappaport Faculty of Medicine, Technion – Israel Institute of Technology, P.O. Box 9649, Bat Galim, Haifa 31096, Israel
1To whom correspondence should be addressed (email debya@tx.technion.ac.il).
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Publisher: Portland Press Ltd
Received:
October 11 2005
Revision Received:
February 22 2006
Accepted:
March 06 2006
Accepted Manuscript online:
March 06 2006
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2006
Biochem J (2006) 397 (1): 213–222.
Article history
Received:
October 11 2005
Revision Received:
February 22 2006
Accepted:
March 06 2006
Accepted Manuscript online:
March 06 2006
Citation
Natalia Volinsky, Anna Gantman, Deborah Yablonski; A Pak- and Pix-dependent branch of the SDF-1α signalling pathway mediates T cell chemotaxis across restrictive barriers. Biochem J 1 July 2006; 397 (1): 213–222. doi: https://doi.org/10.1042/BJ20051655
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