Monitoring glycolytic dynamics in single cells using a fluorescent biosensor for fructose 1,6-bisphosphate.
Monitoring glycolytic dynamics in single cells using a fluorescent biosensor for fructose 1,6-bisphosphate.
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利用荧光生物传感器监测果糖1,6-二磷酸的单细胞糖酵解动力学。
DOI：
10.1073/pnas.2204407119
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发表时间：
2022-08-02
影响因子：
11.1
通讯作者：
中科院分区：
文献类型：
--
作者：
细胞通过调节糖酵解代谢的变化来响应葡萄糖供应的变化或能量需求的波动。葡萄糖转化为丙酮酸的速率对细胞生理学相当重要;然而，在单细胞分辨率下测量该特性从根本上是困难的。糖酵解中间体果糖1，6-二磷酸（FBP）的浓度随流量变化而变化。我们开发了一种荧光生物传感器，利用这种相关性来监测单细胞内糖酵解的快速变化。在胰腺β细胞中使用HYlight以揭示糖酵解调节、动力学和异质性的方面。鉴于糖酵解的保守性和重要性，我们相信这种新的生物传感器将在各种组织和生物体中广泛使用。细胞代谢在空间和时间上受到调节，以确保能量生产与消耗有效匹配。荧光生物传感器是研究代谢的有用工具，因为它们能够以单细胞分辨率实时检测代谢物丰度。为了监测糖酵解，中间体果糖1，6-二磷酸（FBP）是一个特别有用的信号，因为它的浓度与整个途径的通量密切相关。使用GFP插入到枯草芽孢杆菌转录调节因子CggR的配体结合结构域中，我们开发了一种用于FBP的荧光生物传感器，称为HYlight。我们证明，HYlight可以可靠地报告活细胞和组织中糖酵解的实时动态，由各种代谢或药理学扰动驱动，单独或与其他生理相关信号相结合。使用这种传感器，我们发现了以前未知的β细胞糖酵解异质性和动力学方面。
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.
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影响因子：
8.8
作者：
Kondo H;Ratcliffe CDH;Hooper S;Ellis J;MacRae JI;Hennequart M;Dunsby CW;Anderson KI;Sahai E
通讯作者：
Sahai E
影响因子：
7.7
作者：
Civelek, VN;Deeney, JT;Tornheim, K
通讯作者：
Tornheim, K
影响因子：
4.8
作者：
Ardehali, H;Yano, Y;Granner, DK
通讯作者：
Granner, DK
DOI：
10.1074/jbc.m112.414961
发表时间：
2013-04-12
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Lorenz MA;El Azzouny MA;Kennedy RT;Burant CF
通讯作者：
Burant CF
影响因子：
64.8
作者：
ASHCROFT, FM;HARRISON, DE;ASHCROFT, SJH
通讯作者：
ASHCROFT, SJH