Detection of candidate proteins in the indican biosynthetic pathway of Persicaria tinctoria (Polygonum tinctorium) using protein–protein interactions and transcriptome analyses,Phytochemistry

当前位置： X-MOL 学术 › Phytochemistry › 论文详情

Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)

Detection of candidate proteins in the indican biosynthetic pathway of Persicaria tinctoria (Polygonum tinctorium) using protein–protein interactions and transcriptome analyses
Phytochemistry ( IF 3.2 ) Pub Date : 2020-11-01 , DOI: 10.1016/j.phytochem.2020.112507
Shintaro Inoue ₁ , Rihito Morita ₁ , Keiko Kuwata ₂ , Kazuo Ishii ₃ , Yoshiko Minami ₁

Affiliation

Persicaria tinctoria (Polygonum tinctorium) synthesizes indican (indoxyl-β-D-glucoside) as a specialized metabolite. Indican is synthesized in the cytosol of leaf cells from indoxyl and UDP-glucose by the catalysis of indoxyl-β-D-glucoside synthase (PtIGS), then transported into vacuoles. As a portion of PtIGS is found on the microsomal membrane, we assume that it is present on the ER membrane as a large complex involving other indican metabolism-related proteins. Based on this hypothesis, the existence of such a complex was investigated using two separate approaches: a protein-protein interaction assay and transcriptome analysis. We first performed a co-immunoprecipitation using the anti-PtIGS antibody and a pull-down assay using recombinant PtIGS, then identified the candidate proteins through MS/MS analysis. Secondly, we performed a transcriptome analysis to examine the differential gene expression between the first and the second leaves. The expressions of candidate genes detected by protein-protein interaction analyses were collated with transcriptome data and validated by quantitative reverse transcription polymerase chain reaction, showing that the expression of sucrose synthase and cytochrome P450 genes decreased in the second leaves compared with the first leaves. Furthermore, we detected several additional proteins, such as heat shock and cytoskeletal proteins, suggesting that PtIGS may form a large complex, a metabolon.

中文翻译：

使用蛋白质-蛋白质相互作用和转录组分析检测 Persicaria tinctoria (Polygonum tinctorium) 印度生物合成途径中的候选蛋白质

Persicaria tinctoria (Polygonum tinctorium) 合成籼糖（吲哚酚-β-D-葡萄糖苷）作为一种特殊的代谢物。Indican 在叶细胞的细胞质中由吲哚酚和 UDP-葡萄糖在吲哚酚-β-D-葡萄糖苷合酶 (PtIGS) 的催化下合成，然后转运到液泡中。由于 PtIGS 的一部分存在于微粒体膜上，我们假设它作为一个大的复合体存在于 ER 膜上，其中包含其他与印度代谢相关的蛋白质。基于这一假设，使用两种不同的方法研究了这种复合物的存在：蛋白质-蛋白质相互作用分析和转录组分析。我们首先使用抗 PtIGS 抗体进行了免疫共沉淀，并使用重组 PtIGS 进行了下拉测定，然后通过 MS/MS 分析确定了候选蛋白质。第二，我们进行了转录组分析以检查第一片和第二片叶子之间的差异基因表达。通过蛋白质-蛋白质相互作用分析检测到的候选基因的表达与转录组数据进行整理并通过定量逆转录聚合酶链反应进行验证，表明与第一片叶子相比，第二片叶子中蔗糖合酶和细胞色素P450基因的表达降低。此外，我们检测到几种额外的蛋白质，如热休克和细胞骨架蛋白，表明 PtIGS 可能形成一个大的复合物，一种代谢物。通过蛋白质-蛋白质相互作用分析检测到的候选基因的表达与转录组数据进行整理并通过定量逆转录聚合酶链反应进行验证，表明与第一片叶子相比，第二片叶子中蔗糖合酶和细胞色素P450基因的表达降低。此外，我们检测到几种额外的蛋白质，如热休克和细胞骨架蛋白，表明 PtIGS 可能形成一个大的复合物，一种代谢物。通过蛋白质-蛋白质相互作用分析检测到的候选基因的表达与转录组数据进行整理并通过定量逆转录聚合酶链反应进行验证，表明与第一片叶子相比，第二片叶子中蔗糖合酶和细胞色素P450基因的表达降低。此外，我们检测到几种额外的蛋白质，如热休克和细胞骨架蛋白，表明 PtIGS 可能形成一个大的复合物，一种代谢物。

更新日期：2020-11-01

点击分享查看原文

点击收藏

阅读更多本刊最新论文本刊介绍/投稿指南11