Target of rapamycin-signaling modulates starch accumulation via glycogenin phosphorylation status in the unicellular red alga Cyanidioschyzon merolae
Target of rapamycin-signaling modulates starch accumulation via glycogenin phosphorylation status in the unicellular red alga Cyanidioschyzon merolae
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DOI：
10.1111/tpj.14136
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发表时间：
2019-02-01
期刊：
影响因子：
7.2
通讯作者：
Imamura, Sousuke
中科院分区：
文献类型：
--
作者：
Pancha, Imran;Shima, Hiroki;Imamura, Sousuke
雷帕霉素（TOR）信号通路靶点参与多种真核生物淀粉积累；然而，在真核生物中这种现象背后的分子机制尚未阐明。我们报道了TOR在单细胞红藻（Cyanidioschyzon merolae）中淀粉积累的调控机制。经tor特异性抑制剂雷帕霉素灭活后，C. merolae中的淀粉含量比其药物载体二甲亚砜增加了约10倍。然而，我们之前的转录组分析显示，碳水化合物代谢相关基因的表达水平不受雷帕霉素的影响，这表明淀粉积累在转录后水平受到调节。在这项研究中，我们使用液相色谱-串联质谱进行了磷酸化蛋白质组分析，以研究潜在的转录后修饰，并确定了52个候选TOR底物蛋白。在可能的底物中，我们重点研究了CmGLG1的功能，因为它在Ser613残基上的磷酸化在雷帕霉素处理后降低，并且CmGLG1的过表达导致淀粉含量增加4.7倍。CmGLG1与启动淀粉/糖原合成所需的启动蛋白糖原相似，出芽酵母互补实验表明CmGLG1在功能上可以替代糖原。我们发现，与野生型CmGLG1过表达细胞相比，在磷酸化模拟CmGLG1过表达菌株中，Ser613被天冬氨酸取代，淀粉含量降低了约60%。我们的研究结果表明，TOR通过改变C. merolae中CmGLG1 Ser613残基的磷酸化状态来调节淀粉积累。
The target of rapamycin (TOR) signaling pathway is involved in starch accumulation in various eukaryotic organisms; however, the molecular mechanism behind this phenomenon in eukaryotes has not been elucidated. We report a regulatory mechanism of starch accumulation by TOR in the unicellular red alga, Cyanidioschyzon merolae. The starch content in C. merolae after TOR-inactivation by rapamycin, a TOR-specific inhibitor, was increased by approximately 10-fold in comparison with its drug vehicle, dimethyl sulfoxide. However, our previous transcriptome analysis showed that the expression level of genes related to carbohydrate metabolism was unaffected by rapamycin, indicating that starch accumulation is regulated at post-transcriptional levels. In this study, we performed a phosphoproteome analysis using liquid chromatography-tandem mass spectrometry to investigate potential post-transcriptional modifications, and identified 52 proteins as candidate TOR substrates. Among the possible substrates, we focused on the function of CmGLG1, because its phosphorylation at the Ser613 residue was decreased after rapamycin treatment, and overexpression of CmGLG1 resulted in a 4.7-fold higher starch content. CmGLG1 is similar to the priming protein, glycogenin, which is required for the initiation of starch/glycogen synthesis, and a budding yeast complementation assay demonstrated that CmGLG1 can functionally substitute for glycogenin. We found an approximately 60% reduction in the starch content in a phospho-mimicking CmGLG1 overexpression strain, in which Ser613 was substituted with aspartic acid, in comparison with the wild-type CmGLG1 overexpression cells. Our results indicate that TOR modulates starch accumulation by changing the phosphorylation status of the CmGLG1 Ser613 residue in C. merolae.