MacDiarmid Colin W, Taggart Janet, Kubisiak Michael, Eide David J
Department of Nutritional Sciences, University of Wisconsin-Madison, Madison, Wisconsin, USA.
Department of Nutritional Sciences, University of Wisconsin-Madison, Madison, Wisconsin, USA.
J Biol Chem. 2024 Apr;300(4):107147. doi: 10.1016/j.jbc.2024.107147. Epub 2024 Mar 7.
Zinc is required for many critical processes, including intermediary metabolism. In Saccharomyces cerevisiae, the Zap1 activator regulates the transcription of ∼80 genes in response to Zn supply. Some Zap1-regulated genes are Zn transporters that maintain Zn homeostasis, while others mediate adaptive responses that enhance fitness. One adaptive response gene encodes the 2-cysteine peroxiredoxin Tsa1, which is critical to Zn-deficient (ZnD) growth. Depending on its redox state, Tsa1 can function as a peroxidase, a protein chaperone, or a regulatory redox sensor. In a screen for possible Tsa1 regulatory targets, we identified a mutation (cdc19) that partially suppressed the tsa1Δ growth defect. The cdc19 mutation reduced activity of its protein product, pyruvate kinase isozyme 1 (Pyk1), implicating Tsa1 in adapting glycolysis to ZnD conditions. Glycolysis requires activity of the Zn-dependent enzyme fructose-bisphosphate aldolase 1, which was substantially decreased in ZnD cells. We hypothesized that in ZnD tsa1Δ cells, the loss of a compensatory Tsa1 regulatory function causes depletion of glycolytic intermediates and restricts dependent amino acid synthesis pathways, and that the decreased activity of Pyk1 counteracted this depletion by slowing the irreversible conversion of phosphoenolpyruvate to pyruvate. In support of this model, supplementing ZnD tsa1Δ cells with aromatic amino acids improved their growth. Phosphoenolpyruvate supplementation, in contrast, had a much greater effect on growth rate of WT and tsa1Δ ZnD cells, indicating that inefficient glycolysis is a major factor limiting yeast growth. Surprisingly however, this restriction was not primarily due to low fructose-bisphosphate aldolase 1 activity, but instead occurs earlier in glycolysis.
锌参与许多关键过程，包括中间代谢。在酿酒酵母中，Zap1激活剂会根据锌的供应情况调节约80个基因的转录。一些受Zap1调节的基因是维持锌稳态的锌转运蛋白，而其他基因则介导增强适应性的适应性反应。一个适应性反应基因编码2-半胱氨酸过氧化物酶Tsa1，它对缺锌（ZnD）生长至关重要。根据其氧化还原状态，Tsa1可以作为过氧化物酶、蛋白质伴侣或调节性氧化还原传感器发挥作用。在一项寻找可能的Tsa1调节靶点的筛选中，我们鉴定出一个突变（cdc19），该突变部分抑制了tsa1Δ的生长缺陷。cdc19突变降低了其蛋白质产物丙酮酸激酶同工酶1（Pyk1）的活性，这表明Tsa1参与使糖酵解适应ZnD条件。糖酵解需要锌依赖性酶果糖-1,6-二磷酸醛缩酶1的活性，而该酶在ZnD细胞中活性大幅降低。我们推测，在ZnD tsa1Δ细胞中，补偿性Tsa1调节功能的丧失导致糖酵解中间产物耗竭，并限制了相关氨基酸合成途径，而Pyk1活性降低通过减缓磷酸烯醇丙酮酸向丙酮酸的不可逆转化来抵消这种耗竭。为支持该模型，用芳香族氨基酸补充ZnD tsa1Δ细胞可改善其生长。相比之下，补充磷酸烯醇丙酮酸对野生型和tsa1Δ ZnD细胞的生长速率影响更大，表明低效的糖酵解是限制酵母生长的主要因素。然而，令人惊讶的是，这种限制并非主要由于果糖-1,6-二磷酸醛缩酶1活性低，而是发生在糖酵解的更早阶段。
