Regulation of Rac1 and Cdc42 activation in thrombin- and collagen-stimulated CHRF-288-11 cells
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Rac1 and Cdc42 are members of the Rho family of small GTPases and have been shown to promote the formation of lamellipodia and filopodia at the leading edge of motile cells and affect cell migration. In this study the authors have investigated the activation of Rac1 and Cdc42 by thrombin or collagen using the megakaryocytic cell line, CHRF-288-11. Maximal activation of Rac1 by thrombin or collagen was observed at 3 and 1 min, respectively. Similar results were obtained for the activation of Cdc42 by thrombin or collagen. The activation of Rac1 by thrombin and collagen was inhibited by the calmodulin-specific inhibitor, W7. However, W7 had no effect on the activation of Cdc42 by thrombin or collagen. The less potent calmodulin inhibitor, W5, did not have any effect on Rac1 or Cdc42 activation by thrombin or collagen. Transient over-expression of calmodulin in CHRF cells increased the basal and thrombin-mediated activation of Rac1 when compared to control but had no effect on the basal and thrombin-mediated activation of Cdc42 when compared to control. The results suggest that calmodulin regulates the activation of Rac1 in CHRF-288-11 cells in a manner similar to that in the human platelets.
KeywordsRac1 Cdc42 GTPases Thrombin Collagen Calmodulin
The authors wish to thank Dr. Mitsunobu Hoshino (National Center of Neurology and Psychiatry, Tokyo, Japan) for providing the GST-Pak1 plasmid. This study was supported by a grant from the Heart and Stroke Foundation of Manitoba to R.P.B.