Regulation of cell-matrix adhesion dynamics and Rac-1 by integrin linked kinase.
Extracellular matrix (ECM) receptors of the integrin family initiate changes in cell shape and motility by recruiting signaling components that coordinate these events. Integrin-linked kinase (ILK) is one such partner of beta1 integrins that participates in dynamic rearrangement of cell-matrix adhesions and cell spreading by mechanisms that are not well understood. To further elucidate the role of ILK in these events, we engineered a chimeric molecule comprising ILK fused to a membrane-targeted green fluorescent protein (ILK-GFP-F). ILK-GFP-F is highly enriched in cell-matrix adhesions, and its expression in fibroblasts leads to an accumulation of focal adhesions (2-5 microm) and elongated adhesions (>5 microm). ILK-GFP-F enhances cell spreading on fibronectin and induces a constitutive increase in the levels of GTP-bound Rac-1. Conversely, ILK knock-down by siRNA transfection decreases active Rac-1. Endogenous ILK was found to associate with PKL (paxillin kinase linker) and the Rac/Cdc42 guanine nucleotide exchange factor betaPIX. Further, expression of a dominant negative betaPIX mutant reversed the increase in active Rac-1 levels of ILK-GFP-F-expressing cells, thus placing betaPIX in the pathway leading from ILK to Rac-1 activation. However, expression of constitutively active Rac only partially restores the spreading defects of ILK-depleted cells, suggesting that an additional ILK-dependent signal is required for cell spreading.
Boulter
Etienne
E
Institute of Signaling, Developmental Biology and Cancer Research, CNRS-UMR6543, Centre Antoine Lacassagne, 33 Ave. de Valombrose, Nice 06189, France.
Grall
Dominique
D
Cagnol
Sébastien
S
Van Obberghen-Schilling
Ellen
E
D000818:Animals; D001011:Aorta; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D005109:Extracellular Matrix / Q000502:physiology*; D005822:Genetic Vectors; D007700:Kinetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection; D020830:rac1 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D000099200:Scaffold Protein ILK
D017346:Protein Serine-Threonine Kinases; D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D020830:rac1 GTP-Binding Protein; D000099200:Scaffold Protein ILK