Research Abstract
Metabolism has been considered a “solved problem” for many years, a testament to the spectacular work of the 1950’s–70’s as summarized in many wall reference charts. Many thought that now that we understood how cells did their “housekeeping”, we could focus on the more interesting signaling pathways that cancer cells use to evade normal controls on cell division. But during the last decade, a renaissance of interest in metabolism, in the context of cancer, has made it clear that the main function of the all-important oncogenes is actually to “hot-wire” the controls on core metabolic pathways, to enable uncontrolled cell division. And the metabolic state of cells does not just affect acute metabolite availability: levels of specific metabolites can directly influence the epigenetic marks on chromosomes that control gene expression.
Fluorescent biosensors are an important new tool for revealing metabolic behavior. Although we know (for the most part) the metabolic pathways that accomplish biosynthesis and energy production, we are just beginning to learn about how fluxes through these pathways are steered and regulated. Although mass spectroscopy approaches have exquisite chemical specificity, fluorescent biosensors allow us to make real-time movies of metabolic behavior, at the level of individual cells in intact tissue. Particularly in a complex tissue made of heterogeneous cell types – such as brain tissue, or tumor cells in situ – fluorescent imaging can reveal the dynamic behavior of individually targeted cells.
We have developed first-in-class biosensors for key metabolic cofactors – for ATP:ADP ratio and for NADH:NAD+ ratio – as well as a companion sensor for simultaneously monitoring pH. We continue to refine published sensors to make them more practically usable, as well as screening for new sensors. We have also developed methods for high-speed fluorescence lifetime imaging with two-photon laser scanning microscopes, to allow quantitative biosensor imaging of intact (or even in vivo) tissue.
Some of our biological investigations employing the biosensors have focused on the mechanism of metabolic responses to neuronal stimulation, but we have also collaborated with Joan Brugge’s lab on metabolic oscillations in MCF-10A cells, and with Nika Danial’s lab on the altered metabolic behavior in BAD-knockout mice.
