In this Fogarty International Collaboration Award application, we seek to expand the goals set in its parent grant R01GM60523, Interaction of Rho GTPases with regulators and effectors, by examining the mode of regulation and substrate specificity of the Rho GTPase effectors, the p21- activated kinases (PAKs). The Rho family small GTPases Rac1 and Cdc42 regulate coordinated changes to the actin cytoskeleton and gene activation through direct interaction with multiple effector targets. PAK represents a subset of Rac1/Cdc42 effector enzymes that are involved in signaling to both cytoskeleton and nucleus. Recent cellular and genetic studies have shown that PAK plays essential roles in processes including neurite formation and axonal guidance, development of cell polarity and motile responses, and cell proliferation and transformation. To build on the collaborative strengths combining the principal investigator s expertise on the small G-protein-effector interaction and the foreign collaborators experiences in protein kinase and phosphatase enzymology, we set out in this proposal to determine the mode of PAK regulation and to address the specificity issues of PAK signaling pathways.
In specific Aim 1 we will investigate the autophosphorylation mechanism of PAK and examine the dephosphorylation effect of PAK by protein phosphotases to establish an activation/deactivation regulatory cycle.
In specific Aim 2 we will determine the enzymatic specificity of PAK toward a panel of implicated cellular substrates and begin to characterize the PAK-emanated signaling cascades in a case study of one particular downstream event - the Rac-PAK-MLCK-MLC chain reaction that has been implicated in the regulation of smooth muscle contraction. These biochemical approaches are aimed at providing mechanistic insight into the signal transduction modules of the Rho family GTP-binding proteins, malfunction of which may be associated with various human diseases such as neurological disorder, inflammation, and cancer.
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| Wu, Hao; Zheng, Yi; Wang, Zhi-Xin (2003) Evaluation of the catalytic mechanism of the p21-activated protein kinase PAK2. Biochemistry 42:1129-39 |
| Wu, Hao; Wang, Zhi-Xin (2003) The mechanism of p21-activated kinase 2 autoactivation. J Biol Chem 278:41768-78 |
| Zhao, Ju; Wang, Wei-Ning; Tan, Ying-Cai et al. (2002) Effect of Mg(2+) on the kinetics of guanine nucleotide binding and hydrolysis by Cdc42. Biochem Biophys Res Commun 297:653-8 |
| Tan, Ying-Cai; Wu, Hao; Wang, Wei-Ning et al. (2002) Characterization of the interactions between the small GTPase RhoA and its guanine nucleotide exchange factors. Anal Biochem 310:156-62 |
