The response of human leukocytes to chemoattractants generated at inflammatory sites is critical to their role in the innate immune response. Signaling from G protein-coupled chemoattractant receptors to the cellular cytoskeleton involves Rho GTPases and the downstream mediator, p21-activated kinase (PAK). We have identified a novel PAK molecular partner as GEF- H1, a Rho family guanine nucleotide exchange factor (GEF) associated with microtubules. GEF-H1 provides a potential link between Rho GTPase regulation of the actin-myosin cytoskeleton and microtubule dynamics. We will use molecular and genetic approaches to investigate the regulation and biology of PAK and GEF-H1. Critical questions remain about the spatial and temporal regulation of Rho GTPase function in the context of a polarized, chemotactically responding cell. We will apply a novel method of live cell analysis for detecting Rho GTPase activation (FLAIR) to relate the kinetics of Rho GTPase activation to chemotaxis response parameters, to changes in the cellular cytoskeleton, and to biochemical signaling events. The role(s) of PAK and its molecular partner, PIX, as mediators of Rac and Cdc42 in chemotaxis will be investigated, and their mechanisms of action determined using biochemical and genetic approaches. Our ability to achieve these goals relies on our development of FLAIR technology, as well as effective means to introduce protein components into leukocytes and model cell chemotaxis systems. The proposed studies should yield new insights into the regulation of cell motility and the inflammatory response of human leukocytes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM039434-16
Application #
6684134
Study Section
Medical Biochemistry Study Section (MEDB)
Program Officer
Anderson, Richard A
Project Start
1988-02-01
Project End
2005-11-30
Budget Start
2003-12-01
Budget End
2004-11-30
Support Year
16
Fiscal Year
2004
Total Cost
$457,401
Indirect Cost
Name
Scripps Research Institute
Department
Type
DUNS #
781613492
City
La Jolla
State
CA
Country
United States
Zip Code
92037
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Delorme-Walker, Violaine D; Peterson, Jeffrey R; Chernoff, Jonathan et al. (2011) Pak1 regulates focal adhesion strength, myosin IIA distribution, and actin dynamics to optimize cell migration. J Cell Biol 193:1289-303
Zhang, Hui; Sun, Chunxiang; Glogauer, Michael et al. (2009) Human neutrophils coordinate chemotaxis by differential activation of Rac1 and Rac2. J Immunol 183:2718-28
Nalbant, Perihan; Chang, Yuan-Chen; Birkenfeld, Jorg et al. (2009) Guanine nucleotide exchange factor-H1 regulates cell migration via localized activation of RhoA at the leading edge. Mol Biol Cell 20:4070-82
Zhao, Tieming; Nalbant, Perihan; Hoshino, Mikio et al. (2007) Signaling requirements for translocation of P-Rex1, a key Rac2 exchange factor involved in chemoattractant-stimulated human neutrophil function. J Leukoc Biol 81:1127-36
Delorme, Violaine; Machacek, Matthias; DerMardirossian, Celine et al. (2007) Cofilin activity downstream of Pak1 regulates cell protrusion efficiency by organizing lamellipodium and lamella actin networks. Dev Cell 13:646-62
Birkenfeld, Jorg; Nalbant, Perihan; Bohl, Benjamin P et al. (2007) GEF-H1 modulates localized RhoA activation during cytokinesis under the control of mitotic kinases. Dev Cell 12:699-712
Zhao, Tieming; Bokoch, Gary M (2007) Transduction of proteins into intact neutrophils. Methods Mol Biol 412:115-23
Belvindrah, Richard; Nalbant, Perihan; Ding, Sheng et al. (2006) Integrin-linked kinase regulates Bergmann glial differentiation during cerebellar development. Mol Cell Neurosci 33:109-25
Pestonjamasp, Kersi N; Forster, Carol; Sun, Chunxiang et al. (2006) Rac1 links leading edge and uropod events through Rho and myosin activation during chemotaxis. Blood 108:2814-20

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