The role of NAGK cysteine deprotonation in nutrient stress and cancer progression

项目总结/摘要 细胞糖基化的变化支持癌症的生长和进展，但 供应所需的核苷酸-糖前体是资源密集型的。因此，该小组利用打捞 在聚糖降解或细胞外摄取后回收游离糖的途径， 从头合成。关于癌细胞在多大程度上依赖糖补救途径或如何依赖糖补救途径， 这些途径受到调控。最近，补救激酶负责磷酸化N- 乙酰葡糖胺（GlcNAc），通过己糖胺生物合成途径重复使用，N-乙酰-D-葡糖胺激酶 （NAGK），被发现支持肿瘤生长，提供了早期证据，表明癌细胞可能确实严重依赖 打捞上来的本文提供的初步数据表明，NAGK在两个半胱氨酸处去质子化，以响应于 营养限制，这是肿瘤微环境中经常遇到的情况。半胱氨酸去质子化 可以改变蛋白质活性并促进随后的氧化。这种去质子化不仅在NAGK上观察到， 而是对整个细胞中的蛋白质的影响，这可能是以前未被认识到的对压力的适应。半胱氨酸， 特别是在去质子化的硫醇盐状态下，作为小分子共价抑制的现成位点 亲电体因此，广泛增加硫醇盐响应营养限制可能代表一类 在应激肿瘤细胞中，蛋白质对共价抑制的反应比在应激肿瘤细胞中时更敏感。 质子化巯基状态在健康的灌注组织中。本提案中详细阐述的目标将首先描述 半胱氨酸去质子化对NAGK活性的影响及其在肿瘤生长中的作用，然后将集中于鉴定 半胱氨酸质子化状态响应于营养限制以及环境的全球变化 促进这种去质子化的因素。这项工作将特点，无论是机械和广泛的，一部小说 应激反应可能促进肿瘤进展，尽管营养限制，但也可能使 肿瘤更容易受到小分子亲电体的治疗干预。

PROJECT SUMMARY/ ABSTRACT Changes in cellular glycosylation support cancer growth and progression, but the biosynthetic pathways that supply the required nucleotide-sugar precursors are resource intensive. The cell therefore utilizes salvage pathways to recycle free sugars following glycan degradation or extracellular uptake, offsetting the need for their de novo synthesis. Little is known about the extent to which cancer cells rely on sugar salvage pathways or how these pathways are regulated. Recently, the salvage kinase responsible for phosphorylating N- acetylglucosamine (GlcNAc) for reuse by the hexosamine biosynthetic pathway, N-acetyl-D-glucosamine kinase (NAGK), was found to support tumor growth, providing early evidence that cancer cells may indeed rely heavily on salvage. Preliminary data presented here show that NAGK is deprotonated at two cysteines in response to nutrient limitation, a condition frequently encountered in the tumor microenvironment. Cysteine deprotonation can alter protein activity and promote subsequent oxidation. This deprotonation is observed not just on NAGK, but on proteins throughout the cell in what may be a previously unrecognized adaptation to stress. Cysteines, especially in the deprotonated thiolate state, serves as ready sites of covalent inhibition by small molecule electrophiles. Thus, the wide-spread increase in thiolates in response to nutrient limitation may represent a class of proteins that are more responsive to covalent inhibition within stressed tumor cells than when in their protonated thiol state in healthy, perfused tissue. The aims detailed in this proposal will first characterize the effect of cysteine deprotonation on NAGK activity and its role in tumor growth and will then focus on identifying the global changes in cysteine protonation status in response to nutrient limitation as well as the environmental factors that promote this deprotonation. This work will characterize, both mechanistically and broadly, a novel stress response that may promote tumor progression despite nutrient limitation but that may also render the tumor more vulnerable to therapeutic intervention with small molecule electrophiles.