Department of Chemistry, University of Washington, Box 351700, Seattle, WA 98195-7350, USA.
Anal Bioanal Chem. 2011 Nov;401(8):2387-402. doi: 10.1007/s00216-011-4800-2. Epub 2011 Mar 17.
The AMP-activated protein kinase in yeast, Snf1, coordinates expression and activity of numerous intracellular signaling and developmental pathways, including those regulating cellular differentiation, response to stress, meiosis, autophagy, and the diauxic transition. Snf1 phosphorylates metabolic enzymes and transcription factors to change cellular physiology and metabolism. Adr1 and Cat8, transcription factors that activate gene expression after the diauxic transition, are regulated by Snf1; Cat8 through direct phosphorylation and Adr1 by dephosphorylation in a Snf1-dependent manner. Adr1 and Cat8 coordinately regulate numerous genes encoding enzymes of gluconeogenesis, the glyoxylate cycle, β-oxidation of fatty acids, and the utilization of alternative fermentable sugars and nonfermentable substrates. To determine the roles of Adr1, Cat8, and Snf1 in metabolism, two-dimensional gas chromatography coupled to time-of-flight mass spectrometry and liquid chromatography coupled to tandem mass spectrometry were used to identify metabolites whose levels change after the diauxic transition in wild-type-, ADR1-, CAT8-, and SNF1-deficient yeast. A discovery-based approach to data analysis utilized chemometric algorithms to identify, quantify, and compare 63 unique metabolites between wild type, adr1∆, cat8∆, adr1∆cat8∆, and snf1∆ strains. The primary metabolites found to differ were those of gluconeogenesis, the glyoxylate and tricarboxylic acid cycles, and amino acid metabolism. In general, good agreement was observed between the levels of metabolites derived from these pathways and the levels of transcripts from the same strains, suggesting that transcriptional control plays a major role in regulating the levels of metabolites after the diauxic transition.
酵母中的 AMP 激活蛋白激酶 Snf1 协调众多细胞内信号和发育途径的表达和活性，包括那些调节细胞分化、应激反应、减数分裂、自噬和双相转换的途径。Snf1 磷酸化代谢酶和转录因子，以改变细胞生理和代谢。Adr1 和 Cat8 是双相转换后激活基因表达的转录因子，它们受 Snf1 调控；Cat8 通过直接磷酸化，Adr1 通过 Snf1 依赖性去磷酸化进行调控。Adr1 和 Cat8 协同调节许多编码糖异生、乙醛酸循环、脂肪酸β-氧化和替代可发酵糖和不可发酵底物利用的酶的基因。为了确定 Adr1、Cat8 和 Snf1 在代谢中的作用，使用二维气相色谱-飞行时间质谱和液相色谱-串联质谱联用技术，鉴定了在野生型、ADR1、CAT8 和 SNF1 缺陷酵母中双相转换后水平变化的代谢物。基于发现的数据分析方法利用化学计量学算法，在野生型、adr1∆、cat8∆、adr1∆cat8∆和 snf1∆菌株之间识别、定量和比较了 63 种独特的代谢物。发现差异的主要代谢物是糖异生、乙醛酸和三羧酸循环以及氨基酸代谢的代谢物。一般来说，从这些途径获得的代谢物水平与同一菌株的转录物水平之间观察到很好的一致性，这表明转录控制在双相转换后调节代谢物水平方面起着主要作用。
