Gary YellenDepartment of Neurobiology
Harvard Medical School
Warren Alpert Bldg. Room 328
220 Longwood Ave., Boston, MA 02115
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. 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: between neurons and astrocytes, 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 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 parallel. We have developed novel sensors for two key metabolites (ATP and NADH), in order to address specific questions about how metabolism influences neuronal ion channels and can reduce susceptibility to epileptic seizures. They also offer a new window into the dynamic changes of metabolism in many other cell types.
Our lab is working to develop a series of metabolite biosensors, which can report the dynamics of metabolism in individual cells and can be used to complement the powerful tools of mass spec metabolomics. Using structural and functional data on metabolite binding proteins, we are designing and building new fluorescent metabolite sensors by gene synthesis, mutagenesis, and screening/optimization. We use state-of-the-art two-photon scanning microscopy and fluorescence lifetime imaging to image the sensors in living tissue (e.g. brain slices, and in the near future, in vivo recordings of rodent brain).
We also study how metabolic manipulation, through either dietary change or genetic alteration, can alter brain excitability, using mouse models and brain slice electrophysiology.
Ma, W., Berg, J., Yellen, G. (2007) Ketogenic diet metabolites reduce firing in central neurons by opening KATP channels. J Neurosci. 27:3618-25.
Berg, J., Hung, Y.P., Yellen, G. (2009) A genetically encoded fluorescent reporter of ATP:ADP ratio. Nature Methods 6:161-6.
Tanner, G.R., Lutas, A., Martínez-François, J.R., Yellen, G. (2011) Single KATP channel opening in response to action potential firing in mouse dentate granule neurons. J Neurosci. 31:8689-96.
Hung, Y.P., Albeck, J.G, Tantama, M., Yellen, G. (2011) Imaging cytosolic NADH-NAD+ redox state with a genetically encoded fluorescent biosensor. Cell Metabolism 14:545-54.
Giménez-Cassina, A., Martínez-François, J.R., Fisher, J.K., Szlyk, B., Polak, K., Wiwczar, J., Tanner, G.R., Lutas, A., Yellen, G., Danial, N.N. (2012) BAD-dependent regulation of fuel metabolism and KATP channel activity confers resistance to epileptic seizures. Neuron 74:719-30.
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