期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：31
DOI：10.1073/pnas.2204407119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance
Cells respond to shifts in glucose supply or fluctuations in energetic demand through regulated changes in glycolytic metabolism. The rate at which glucose is converted to pyruvate is of considerable importance to cellular physiology; however, measuring this property at single-cell resolution is fundamentally difficult. The glycolytic intermediate fructose 1,6-bisphosphate (FBP) exhibits large changes in concentration that track with changes in flux. We developed a fluorescent biosensor for FBP exploiting this correlation to monitor rapid changes in glycolysis within single cells. HYlight was used in pancreatic β-cells to uncover aspects of glycolytic regulation, dynamics and heterogeneity. Given the conservation and importance of glycolysis, we believe this new biosensor will be broadly useful in a variety of tissues and organisms.
Cellular metabolism is regulated over space and time to ensure that energy production is efficiently matched with consumption. Fluorescent biosensors are useful tools for studying metabolism as they enable real-time detection of metabolite abundance with single-cell resolution. For monitoring glycolysis, the intermediate fructose 1,6-bisphosphate (FBP) is a particularly informative signal as its concentration is strongly correlated with flux through the whole pathway. Using GFP insertion into the ligand-binding domain of the
Bacillus subtilis transcriptional regulator CggR, we developed a fluorescent biosensor for FBP termed HYlight. We demonstrate that HYlight can reliably report the real-time dynamics of glycolysis in living cells and tissues, driven by various metabolic or pharmacological perturbations, alone or in combination with other physiologically relevant signals. Using this sensor, we uncovered previously unknown aspects of β-cell glycolytic heterogeneity and dynamics.
