To appreciate how cells migrate, establish polarity, and adopt cell shape is fundamental to understanding cancer biology. Almost one percent of the human proteome is dedicated to Rho GTPase signaling, which regulates key aspects of each of these processes. It is well known that Rho GTPases are """"""""signaling switches"""""""" that are turned """"""""on"""""""" or """"""""off"""""""" by GEF (Guanine nucleotide Exchange Factors) and GAP (GTPase Activating Proteins) proteins. What is not known are the molecular mechanisms that determine Rho GEF and GAP specificity. Because GEFs and GAPs regulate Rho-GTPases in specific cellular contexts, they are promising candidates for future therapeutic interventions to modulate GTPase activity in different types of cancers. This proposal develops an innovative and integrated approach to map the architecture of Rho GTPase signaling in vivo. Multiple independent methods will be used, including novel proteomic screens using cellular proteins with unnatural amino acids as well as high throughput binding assays and informatics- based experiments. We have recently established this approach in our laboratory and our preliminary data validates our rationale that this approach will uncover novel links between Rho GTPase signaling and cancer biology. At the conclusion of this pilot project we expect to demonstrate that this approach can work on a larger scale to map interaction networks for regulators of this pathway. These results will fundamentally advance our understanding of how Rho GTPases are modulated to regulate processes such as migration, polarity, and cell morphology. It can also be expected to provide the basic information necessary for the future development of therapeutic strategies to alter cancer outcomes.

Public Health Relevance

Because GEFs and GAPs regulate Rho-GTPases in specific cellular contexts, they are promising candidates for future therapeutic interventions to modulate GTPase activity in different types of cancers. This proposal develops an innovative and integrated approach that uses emerging technologies to map the architecture of Rho GTPase signaling in vivo. Multiple independent methods will be used, including novel proteomic screens using cellular proteins with unnatural amino acids as well as high throughput binding assays and informatics-based experiments.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21CA140030-02
Application #
7896620
Study Section
Special Emphasis Panel (ZCA1-SRLB-V (M1))
Program Officer
Knowlton, John R
Project Start
2009-08-01
Project End
2012-01-31
Budget Start
2010-08-01
Budget End
2012-01-31
Support Year
2
Fiscal Year
2010
Total Cost
$171,600
Indirect Cost
Name
Duke University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
044387793
City
Durham
State
NC
Country
United States
Zip Code
27705
Okada, Hirokazu; Uezu, Akiyoshi; Soderblom, Erik J et al. (2012) Peptide array X-linking (PAX): a new peptide-protein identification approach. PLoS One 7:e37035
Uezu, Akiyoshi; Okada, Hirokazu; Murakoshi, Hideji et al. (2012) Modified SH2 domain to phototrap and identify phosphotyrosine proteins from subcellular sites within cells. Proc Natl Acad Sci U S A 109:E2929-38
Okada, Hirokazu; Uezu, Akiyoshi; Mason, Frank M et al. (2011) SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes. Sci Signal 4:rs13