The S. cerevisiae ISR1 gene encodes a putative kinase with no ascribed function. Here, we show that Isr1 acts as a negative regulator of the highly-conserved hexosamine biosynthesis pathway (HBP), which converts glucose into uridine diphosphate N-acetylglucosamine (UDP-GlcNAc), the carbohydrate precursor to protein glycosylation, GPI-anchor formation, and chitin biosynthesis. Overexpression of ISR1 is lethal and, at lower levels, causes sensitivity to tunicamycin and resistance to calcofluor white, implying impaired protein glycosylation and reduced chitin deposition. Gfa1 is the first enzyme in the HBP and is conserved from bacteria and yeast to humans. The lethality caused by ISR1 overexpression is rescued by co-overexpression of GFA1 or exogenous glucosamine, which bypasses GFA1’s essential function. Gfa1 is phosphorylated in an Isr1-dependent fashion and mutation of Isr1-dependent sites ameliorates the lethality associated with ISR1 overexpression. Isr1 contains a phosphodegron that is phosphorylated by Pho85 and subsequently ubiquitinated by the SCF-Cdc4 complex, largely confining Isr1 protein levels to the time of bud emergence. Mutation of this phosphodegron stabilizes Isr1 and recapitulates the overexpression phenotypes. As Pho85 is a cell cycle and nutrient responsive kinase, this tight regulation of Isr1 may serve to dynamically regulate flux through the HBP and modulate how the cell’s energy resources are converted into structural carbohydrates in response to changing cellular needs.

S 。酿酒酵母 ISR1基因编码一种推定的激酶，没有特定的功能。在这里，我们证明 Isr1 是高度保守的己糖胺生物合成途径 (HBP) 的负调节因子，该途径将葡萄糖转化为尿苷二磷酸 N-乙酰氨基葡萄糖 (UDP-GlcNAc)，这是蛋白质糖基化、GPI 锚定形成的碳水化合物前体，和甲壳素生物合成。 ISR1的过度表达是致命的，并且在较低水平下会导致对衣霉素的敏感性和对 calcoflor White 的抗性，这意味着蛋白质糖基化受损和几丁质沉积减少。 Gfa1 是 HBP 中的第一个酶，从细菌、酵母到人类都是保守的。 ISR1过度表达引起的致死性可以通过GFA1或外源葡萄糖胺的共过度表达来挽救，这绕过了GFA1 的基本功能。 Gfa1 以 Isr1 依赖性方式磷酸化，Isr1 依赖性位点的突变可改善与ISR1过表达相关的致死率。 Isr1 含有一个磷酸化子，该磷酸化子被 Pho85 磷酸化，随后被 SCF-Cdc4 复合物泛素化，很大程度上将 Isr1 蛋白水平限制在芽出现时。该磷酸降解子的突变可稳定 Isr1 并重现过度表达表型。由于 Pho85 是一种细胞周期和营养响应激酶，Isr1 的这种严格调节可能有助于动态调节通过 HBP 的通量，并调节细胞的能量资源如何转化为结构碳水化合物，以响应不断变化的细胞需求。