PhyLife, Institute of Biochemistry and Molecular Biology, University of Southern Denmark, Odense M, Denmark.
Yeast. 2024 Aug;41(8):486-498. doi: 10.1002/yea.3972. Epub 2024 Jun 21.
We have investigated the interplay between glycolytic oscillations and intracellular concentration in the yeast Saccharomyces cerevisiae. Intracellular concentration was measured using the fluorophore potassium-binding benzofuranisophthalate (PBFI). We found that is an essential ion for the occurrence of glycolytic oscillations and that intracellular concentration oscillates synchronously with other variables such as nicotinamide adenine dinucleotide hydride (NADH), intracellular adenosine triphosphate (ATP), and mitochondrial membrane potential. We also investigated if glycolysis and intracellular concentration oscillate in a number of yeast strains with mutations in transporters in the plasma membrane, mitochondrial membrane and in the vacuolar membrane. Most of these strains are still capable of showing glycolytic oscillations, but two strains are not: (i) a strain with a deletion in the mitochondrial Mdm38p transporter and (ii) a strain with deletion of the late endosomal Nhx1p ( ) transporter. In these two mutant strains intracellular concentration seems to be low, indicating that the two transporters may be involved in transport of into the cytosol. In the strain, Mdm38p oscillations in glycolysis could be restored by addition of the exchange ionophore nigericin. Furthermore, in two nonoscillating mutant strains with a defective V-ATPase and deletion of the Arp1p protein the intracellular is relatively high, suggesting that the V-ATPase is essential for transport of out of the cytosol and that the cytoskeleton may be involved in binding to reduce the concentration of free ion in the cytosol. Analyses of the time series of oscillations of NADH, ATP, mitochondrial membrane potential, and potassium concentration using data-driven modeling corroborate the conjecture that ion is essential for the emergence of oscillations and support the experimental findings using mutant strains.
我们研究了糖酵解振荡和酵母细胞内浓度之间的相互作用。使用荧光染料钾结合苯并呋喃异苯并二氢吡喃(PBFI)测量细胞内浓度。我们发现是发生糖酵解振荡所必需的离子,并且细胞内浓度与其他变量(如烟酰胺腺嘌呤二核苷酸氢化物(NADH)、细胞内三磷酸腺苷(ATP)和线粒体膜电位)同步振荡。我们还研究了在质膜、线粒体膜和液泡膜中的转运体突变的酵母菌株中,糖酵解和细胞内浓度是否振荡。大多数这些菌株仍然能够显示糖酵解振荡,但有两个菌株不能:(i)线粒体 Mdm38p 转运体缺失的菌株和(ii)晚期内体 Nhx1p ( )转运体缺失的菌株。在这两个突变菌株中,细胞内浓度似乎较低,表明这两种转运体可能参与将 转运到细胞质中。在该菌株中,通过添加 交换离子载体 Nigericin,可以恢复 Mdm38p 在糖酵解中的振荡。此外,在具有缺陷的 V-ATPase和 Arp1p 蛋白缺失的两个非振荡突变菌株中,细胞内 相对较高,表明 V-ATPase对于将 从细胞质中转运出来是必需的,并且细胞骨架可能参与结合以降低细胞质中游离离子的浓度。使用基于数据的建模对 NADH、ATP、线粒体膜电位和钾浓度的振荡时间序列进行分析,证实了 离子对于振荡出现是必需的猜想,并支持使用突变菌株的实验结果。