Quantitative mass spectrometry-based multiplexing compares the abundance of 5000 S-cerevisiae proteins across 10 carbon sources
Quantitative mass spectrometry-based multiplexing compares the abundance of 5000 S-cerevisiae proteins across 10 carbon sources
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DOI：
10.1016/j.jprot.2016.07.005
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发表时间：
2016-10-04
影响因子：
3.3
通讯作者：
Gygi, Steven P.
中科院分区：
文献类型：
--
作者：
Paulo, Joao A.;O'Connell, Jeremy D.;Gygi, Steven P.
芽殖酵母酿酒酵母是研究生物过程的模型系统。已知细胞事件由于碳源的变化而失调。然而，其全面的蛋白质组学改变尚未得到充分研究。在这里，我们研究了S.在酿酒酵母中，由于酵母从葡萄糖适应九种不同的碳源-麦芽糖、海藻糖、果糖、蔗糖、甘油、乙酸盐、丙酮酸盐、乳酸和油酸盐，因此酵母可以从葡萄糖中产生葡萄糖。基于同位素标签的质谱技术处于全球蛋白质组学研究的最前沿。因此，我们使用TMT 10-plex策略在单个实验中研究多种生长条件。在Orbitrap Fusion Lumos质谱仪上的SPS-MS 3方法能够在1%蛋白质水平FDR下对10种碳源中的5000多种酵母蛋白质进行定量。平均而言，在果糖和蔗糖中培养的酵母的蛋白质组与在葡萄糖中培养的蛋白质组的偏差最小。正如预期的那样，基因本体分类揭示了蛋白质丰度的主要变化发生在代谢途径和线粒体蛋白。我们的方案为进一步研究碳源诱导的蛋白质改变奠定了基础。此外，这些数据提供了一个假设产生的资源，为未来的研究，旨在调查的特点和uncharacterized基因。在酿酒酵母中，酵母从葡萄糖适应九种不同的碳源-麦芽糖、海藻糖、果糖、蔗糖、甘油、乙酸盐、丙酮酸盐、乳酸和油酸盐。SPS-MS 3 TMT 10 plex分析用于定量蛋白质组学分析。我们展示了一种技术，可以量化超过5000种酵母蛋白质，这是迄今为止S.酿酒酵母，在单一实验中跨越10种生长条件。正如预期的那样，基因本体分类的蛋白质与丰度发生重大变化的代谢途径和线粒体蛋白，反映了代谢压力的程度时，细胞机械从葡萄糖上的生长转移到替代碳源。我们的方案奠定了基础，为进一步研究碳源诱导的蛋白质改变。提高覆盖深度-测量超过5000种蛋白质的丰度变化-增加了我们对模型系统S中难以研究的基因的理解。酿酒酵母和同源人类细胞生物学。我们提交了这个高度综合的数据集作为一个假设生成资源，有针对性的研究未表征的基因。(C)© 2016 Elsevier B. V.版权所有。
The budding yeast Saccharomyces cerevisiae is a model system for investigating biological processes. Cellular events are known to be dysregulated due to shifts in carbon sources. However, the comprehensive proteomic alterations thereof have not been fully investigated. Here we examined proteomic alterations in S. cerevisiae due to the adaptation of yeast from glucose to nine different carbon sources - maltose, trehalose, fructose, sucrose, glycerol, acetate, pyruvate, lactic acid, and oleate. Isobaric tag-based mass spectrometry techniques are at the forefront of global proteomic investigations. As such, we used a TMT10-plex strategy to study multiple growth conditions in a single experiment. The SPS-MS3 method on an Orbitrap Fusion Lumos mass spectrometer enabled the quantification of over 5000 yeast proteins across ten carbon sources at a 1% protein-level FDR. On average, the proteomes of yeast cultured in fructose and sucrose deviated the least from those cultured in glucose. As expected, gene ontology classification revealed the major alteration in protein abundances occurred in metabolic pathways and mitochondrial proteins. Our protocol lays the groundwork for further investigation of carbon source induced protein alterations. Additionally, these data offer a hypothesis-generating resource for future studies aiming to investigate both characterized and uncharacterized genes.Biological significance: We investigate the proteomic alterations in S. cerevisiae resulting from adaptation of yeast from glucose to nine different carbon sources - maltose, trehalose, fructose, sucrose, glycerol, acetate, pyruvate, lactic acid, and oleate. SPS-MS3 TMT10plex analysis is used for quantitative proteomic analysis. We showcase a technique that allows the quantification of over 5000 yeast proteins, the highest number to date in S. cerevisiae, across 10 growth conditions in a single experiment. As expected, gene ontology classification of proteins with the major alterations in abundances occurred in metabolic pathways and mitochondrial proteins, reflecting the degree of metabolic stress when cellular machinery shifts from growth on glucose to an alternative carbon source. Our protocol lays the groundwork for further investigation of carbon source-induced protein alterations. Improving depth of coverage - measuring abundance changes of over 5000 proteins - increases our understanding of difficult-to-study genes in the model system S. cerevisiae and by homology human cell biology. We submit this highly comprehensive dataset as a hypothesis generating resource for targeted studies on uncharacterized genes. (C) 2016 Elsevier B.V. All rights reserved.