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iTRAQ 标记的全局磷酸化蛋白质组学揭示了不同基因型菌株之间的新型分子差异。

iTRAQ-Based Global Phosphoproteomics Reveals Novel Molecular Differences Between Strains of Different Genotypes.

机构信息

State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, China.

College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, China.

出版信息

Front Cell Infect Microbiol. 2019 Aug 23;9:307. doi: 10.3389/fcimb.2019.00307. eCollection 2019.

DOI:10.3389/fcimb.2019.00307
PMID:31508380
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6716450/
Abstract

To gain insights into differences in the virulence among strains at the post-translational level, we conducted a quantitative analysis of the phosphoproteome profile of strains belonging to three different genotypes. Phosphopeptides from three strains, type I (RH strain), type II (PRU strain) and ToxoDB#9 (PYS strain), were enriched by titanium dioxide (TiO2) affinity chromatography and quantified using iTRAQ technology. A total of 1,441 phosphopeptides, 1,250 phosphorylation sites and 759 phosphoproteins were detected. In addition, 392, 298, and 436 differentially expressed phosphoproteins (DEPs) were identified in RH strain when comparing RH/PRU strains, in PRU strain when comparing PRU/PYS strains, and in PYS strain when comparing PYS/RH strains, respectively. Functional characterization of the DEPs using GO, KEGG, and STRING analyses revealed marked differences between the three strains. kinase substrate motif analysis of the DEPs revealed three (RxxS, SxxE, and SxxxE), three (RxxS, SxxE, and SP), and five (SxxE, SP, SxE, LxRxxS, and RxxS) motifs in RH strain when comparing RH/PRU strains, in PRU strain when comparing PRU/PYS, and in PYS strain when comparing PYS/RH strains, respectively. This suggests that multiple overrepresented protein kinases including PKA, PKG, CKII, IKK, and MAPK could be involved in such a difference between strains. Kinase associated network analysis showed that ROP5, ROP16, and cell-cycle-associated protein kinase CDK were the most connected kinase peptides. Our data reveal significant changes in the abundance of phosphoproteins between genotypes, which explain some of the mechanisms that contribute to the virulence heterogeneity of this parasite.

摘要

为了深入了解不同基因型间菌株在翻译后水平上毒力的差异，我们对属于三种不同基因型的菌株的磷酸化蛋白质组谱进行了定量分析。采用 TiO2 亲和层析法从三种菌株（I 型（RH 株）、II 型（PRU 株）和 ToxoDB#9（PYS 株））中富集磷酸肽，并使用 iTRAQ 技术进行定量。共检测到 1441 个磷酸肽、1250 个磷酸化位点和 759 个磷酸化蛋白质。此外，在 RH 株与 RH/PRU 株比较时、在 PRU 株与 PRU/PYS 株比较时和在 PYS 株与 PYS/RH 株比较时，分别鉴定到 392、298 和 436 个差异表达的磷酸化蛋白质（DEPs）。通过 GO、KEGG 和 STRING 分析对 DEPs 进行功能特征描述，揭示了这三种菌株之间的明显差异。对 DEPs 的激酶底物基序分析表明，在 RH 株与 RH/PRU 株比较时、在 PRU 株与 PRU/PYS 株比较时和在 PYS 株与 PYS/RH 株比较时，分别有三个（RxxS、SxxE 和 SxxxE）、三个（RxxS、SxxE 和 SP）和五个（SxxE、SP、SxE、LxRxxS 和 RxxS）基序。这表明多个过度表达的蛋白激酶，包括 PKA、PKG、CKII、IKK 和 MAPK，可能参与了这种菌株间的差异。激酶相关网络分析表明，ROP5、ROP16 和细胞周期相关蛋白激酶 CDK 是最具连接性的激酶肽。我们的数据揭示了不同基因型间磷酸化蛋白质丰度的显著变化，这解释了该寄生虫毒力异质性的一些机制。

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