Metabolic pathways provide essential energy and building blocks for the function of all cells, and dysregulation of these pathways is a central feature of cancer, diabetes, and obesity, which kill or disable millions of Americans every year. The components of core metabolic pathways such as glycolysis have been very well understood for decades, but there are still major gaps in our understanding of their integrated behavior and regulation in the context of living cells. A major challenge to understanding normal metabolism and its dysregulation in human disease is that metabolic behavior can vary dramatically from cell to cell, and over time within a single cell. For example, metabolic state can differ radically between neighboring cell types in a tissue, creating a functional segregation that is important for overall tissue function, or between a single metastatic cancer cell and the surrounding normal cells. Such spatial differences as well as dynamic changes in metabolism within a single cell are invisible to the usual biochemical methods or even modern metabolomic methods, which require disruption of the living cell and homogenization of tissue. Fluorescent sensors of metabolism, engineered by combining fluorescent proteins with metabolite binding proteins, can address this challenge by enabling us to monitor key metabolites in real time, in single living cells, or in hundreds of cells in paralle. We recently piloted the development of novel sensors for two key metabolites (ATP and NADH), in order to address specific neurobiological questions about how metabolism influences neuronal ion channels and can reduce susceptibility to epileptic seizures. But our preliminary results with these sensors have underscored the general problem of cell heterogeneity as well as the need for a much larger toolkit of fluorescent metabolite sensors. We propose to develop a suite of novel sensors for key metabolites in order to address fundamental questions of cellular me

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
NIH Director’s Pioneer Award (NDPA) (DP1)
Project #
1DP1EB016985-01
Application #
8341600
Study Section
Special Emphasis Panel (ZGM1-NDPA-A (01))
Program Officer
Conroy, Richard
Project Start
2012-09-30
Project End
2017-07-31
Budget Start
2012-09-30
Budget End
2013-07-31
Support Year
1
Fiscal Year
2012
Total Cost
$845,416
Indirect Cost
$345,416
Name
Harvard University
Department
Biology
Type
Schools of Medicine
DUNS #
047006379
City
Boston
State
MA
Country
United States
Zip Code
02115
Chen, Yao; Saulnier, Jessica L; Yellen, Gary et al. (2014) A PKA activity sensor for quantitative analysis of endogenous GPCR signaling via 2-photon FRET-FLIM imaging. Front Pharmacol 5:56
Tantama, Mathew; Martinez-Francois, Juan Ramon; Mongeon, Rebecca et al. (2013) Imaging energy status in live cells with a fluorescent biosensor of the intracellular ATP-to-ADP ratio. Nat Commun 4:2550