Cell Cycle Group, Cancer Epigenetics and Biology Program, Institut d'Investigacions Biomèdica de Bellvitge (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.
IDIBELL Proteomics Unit, Institut d'Investigacions Biomèdica de Bellvitge, L'Hospitalet de Llobregat, Barcelona, Spain.
Gigascience. 2018 May 1;7(5). doi: 10.1093/gigascience/giy047.
Protein phosphatase 2A (PP2A) is a family of conserved serine/threonine phosphatases involved in several essential aspects of cell growth and proliferation. PP2ACdc55 phosphatase has been extensively related to cell cycle events in budding yeast; however, few PP2ACdc55 substrates have been identified. Here, we performed a quantitative mass spectrometry approach to reveal new substrates of PP2ACdc55 phosphatase and new PP2A-related processes in mitotic arrested cells.
We identified 62 statistically significant PP2ACdc55 substrates involved mainly in actin-cytoskeleton organization. In addition, we validated new PP2ACdc55 substrates such as Slk19 and Lte1, involved in early and late anaphase pathways, and Zeo1, a component of the cell wall integrity pathway. Finally, we constructed docking models of Cdc55 and its substrate Mob1. We found that the predominant interface on Cdc55 is mediated by a protruding loop consisting of residues 84-90, thus highlighting the relevance of these aminoacids for substrate interaction.
We used phosphoproteomics of Cdc55-deficient cells to uncover new PP2ACdc55 substrates and functions in mitosis. As expected, several hyperphosphorylated proteins corresponded to Cdk1-dependent substrates, although other kinases' consensus motifs were also enriched in our dataset, suggesting that PP2ACdc55 counteracts and regulates other kinases distinct from Cdk1. Indeed, Pkc1 emerged as a novel node of PP2ACdc55 regulation, highlighting a major role of PP2ACdc55 in actin cytoskeleton and cytokinesis, gene ontology terms significantly enriched in the PP2ACdc55-dependent phosphoproteome.
蛋白磷酸酶 2A（PP2A）是一个家族的保守丝氨酸/苏氨酸磷酸酶，参与细胞生长和增殖的几个基本方面。PP2ACdc55 磷酸酶与芽殖酵母的细胞周期事件广泛相关；然而，已经鉴定出的 PP2ACdc55 底物很少。在这里，我们采用定量质谱方法揭示了 PP2ACdc55 磷酸酶的新底物和有丝分裂期细胞中与 PP2A 相关的新过程。
我们鉴定了 62 个具有统计学意义的 PP2ACdc55 底物，主要涉及肌动蛋白细胞骨架组织。此外，我们验证了新的 PP2ACdc55 底物，如 Slk19 和 Lte1，它们参与早后期和后期有丝分裂途径，以及 Zeo1，细胞壁完整性途径的一个组成部分。最后，我们构建了 Cdc55 及其底物 Mob1 的对接模型。我们发现 Cdc55 的主要界面是由一个由残基 84-90 组成的突出环介导的，从而突出了这些氨基酸对底物相互作用的重要性。
我们使用 Cdc55 缺陷细胞的磷酸蛋白质组学来揭示有丝分裂过程中 PP2ACdc55 的新底物和功能。正如预期的那样，几个过磷酸化的蛋白质对应于 Cdk1 依赖性底物，尽管我们的数据集中还富集了其他激酶的共识基序，这表明 PP2ACdc55 与 Cdk1 不同的其他激酶相互作用并调节它们。事实上，Pkc1 作为 PP2ACdc55 调节的一个新节点出现，突出了 PP2ACdc55 在肌动蛋白细胞骨架和胞质分裂中的主要作用，这是 PP2ACdc55 依赖性磷酸蛋白质组中显著富集的基因本体术语。
Zotero 插件
