We will make fully operational a continuing pilot collaboration between the labs of Dr. Jessica P. Houston (NMSU) and Dr. Roger Brent (FHCRC). Our proposed work builds on complementary strengths in the two labs: instrumentation engineering and signal analysis in the Houston lab, protein engineering and genetics in the Brent lab. In particular, the work builds on "frequency domain" methods to determine fluorescence lifetimes in microscopy, and on significantly more advanced developments in "frequency domain" methods for flow cytometry, for which Dr. Houston is emerging as a leader. Dysfunctions in cell signaling leading to inappropriate cell proliferation contribute to most cancers. Although much is known about cell signaling, key questions, including the causes and consequences of cell-to-cell variation in signaling and response, remain unanswered. Here, we will use fluorescent lifetime methods to extend the power of single cell assays to quantify key events in cell signaling in yeast (the "development platform") and in mammalian cells. These measurements depend on the use of Green Fluorescent Protein (GFP) derivatives fused to the proteins that make up the different cell signaling pathways to quantify events such as recruitment of a signaling complex to the cell membrane. During the next three years, we will develop our methods, to address these questions and to allow wider applications. In particular, we will continue to improve our instrumentation, use it to develop better GFP derivatives and signaling reporters, and use the improved methods to understand the causes of cell-to-cell variation in a yeast signaling system. We will also use these methods to develop high-signal reporters that allow quantification of cell signaling in clonal, mammalian cell lines using flow cytometry. Successful work will merit future funding. It should also increase the visibility of the Houston lab and help recruit new students and researchers from underrepresented ethnic minorities into an active area of international scientific research.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Specialized Center--Cooperative Agreements (U54)
Project #
2U54CA132381-06A1
Application #
8744899
Study Section
Special Emphasis Panel (ZCA1-SRLB-Y (O1))
Project Start
2013-09-25
Project End
2018-08-31
Budget Start
2013-09-25
Budget End
2014-08-31
Support Year
6
Fiscal Year
2013
Total Cost
$171,068
Indirect Cost
$57,666
Name
Fred Hutchinson Cancer Research Center
Department
Type
DUNS #
078200995
City
Seattle
State
WA
Country
United States
Zip Code
98109
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