Systems Biology Lab/AIMMS, Vrije Universiteit Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands.
ACS Sens. 2020 Mar 27;5(3):814-822. doi: 10.1021/acssensors.9b02475. Epub 2020 Mar 3.
Adenosine 5-triphosphate (ATP) is the main free energy carrier in metabolism. In budding yeast, shifts to glucose-rich conditions cause dynamic changes in ATP levels, but it is unclear how heterogeneous these dynamics are at a single-cell level. Furthermore, pH also changes and affects readout of fluorescence-based biosensors for single-cell measurements. To measure ATP changes reliably in single yeast cells, we developed yAT1.03, an adapted version of the AT1.03 ATP biosensor, that is pH-insensitive. We show that pregrowth conditions largely affect ATP dynamics during transitions. Moreover, single-cell analyses showed a large variety in ATP responses, which implies large differences of glycolytic startup between individual cells. We found three clusters of dynamic responses, and we show that a small subpopulation of wild-type cells reached an imbalanced state during glycolytic startup, characterized by low ATP levels. These results confirm the need for new tools to study dynamic responses of individual cells in dynamic environments.
三磷酸腺苷（ATP）是代谢中主要的自由能载体。在出芽酵母中，转移到富含葡萄糖的条件会引起 ATP 水平的动态变化，但在单细胞水平上这些动态变化的异质性如何尚不清楚。此外，pH 值也会发生变化，并影响基于荧光的单细胞测量生物传感器的读出。为了在单个酵母细胞中可靠地测量 ATP 变化，我们开发了 yAT1.03，这是 AT1.03 ATP 生物传感器的适应版本，对 pH 值不敏感。我们表明，预生长条件在转变过程中很大程度上影响了 ATP 的动态变化。此外，单细胞分析显示 ATP 反应存在很大的多样性，这意味着单个细胞之间的糖酵解起始存在很大差异。我们发现了三种动态反应簇，我们表明，在糖酵解起始期间，野生型细胞的一小部分亚群会达到失衡状态，其特征是 ATP 水平低。这些结果证实了需要新的工具来研究动态环境中单个细胞的动态响应。
