Molecular characterization of Clonorchis sinensis secretory myoglobin: Delineating its role in anti-oxidative survival

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• 作者：Mengyu Ren (1) (2)
  Lei He (1) (2)
  Yan Huang (1) (2)
  Qiang Mao (1) (2)
  Shan Li (1) (2)
  Honglin Qu (1) (2)
  Meng Bian (1) (2)
  Pei Liang (3)
  Xueqing Chen (1) (2)
  Jinsi Ling (1) (2)
  Tingjing Chen (1) (2)
  Chi Liang (1) (2)
  Xiaoyun Wang (1) (2)
  Xuerong Li (1) (2)
  Xinbing Yu (1) (2)
  
  1. Department of Parasitology ; Zhongshan School of Medicine ; Sun Yat-sen University ; 74 Zhongshan 2nd Road ; Guangzhou ; Guangdong ; 510080 ; China
  2. Key Laboratory for Tropical Diseases Control (Sun Yat-sen University) ; Ministry of Education ; 74 Zhongshan 2nd Road ; Guangzhou ; Guangdong ; 510080 ; China
  3. Department of Pathogen Biology ; Hainan Medical College ; Haikou ; Hainan ; 571199 ; China
  
• 关键词：Myoglobin ; Clonorchis sinensis ; Point mutation ; Hydrogen peroxide ; RAW264.7 ; Peroxidase activity
• 刊名：Parasites & Vectors
• 出版年：2014
• 出版时间：December 2014
• 年：2014
• 卷：7
• 期：1
• 全文大小：764 KB
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• 刊物主题：Parasitology; Infectious Diseases; Tropical Medicine; Entomology;
• 出版者：BioMed Central
• ISSN：1756-3305

文摘

Background Clonorchiasis is a globally important, neglected food-borne disease caused by Clonorchis sinensis (C. sinensis), and it is highly related to cholangiocarcinoma and hepatocellular carcinoma. Increased molecular evidence has strongly suggested that the adult worm of C. sinensis continuously releases excretory-secretory proteins (ESPs), which play important roles in the parasite-host interactions, to establish successful infection and ensure its own survival. Myoglobin, a hemoprotein, is present in high concentrations in trematodes and ESPs. To further understand the biological function of CsMb and its putative roles in the interactions of C. sinensis with its host, we explored the molecular characterization of CsMb in this paper. Methods We expressed CsMb and its mutants in E. coli BL21 and identified its molecular characteristics using bioinformatics analysis and experimental approaches. Reverse transcription PCR analysis was used to measure myoglobin transcripts of C. sinensis with different culture conditions. The peroxidase activity of CsMb was confirmed by spectrophotometry. We co-cultured RAW264.7 cells with recombinant CsMb (rCsMb), and we then measured the production of hydrogen peroxide (H2O2) and nitric oxide (NO) in addition to the mRNA levels of inducible nitric oxide synthase (iNOS), Cu-Zn superoxide dismutase (SOD1) and Mn superoxide dismutase (SOD2) in activated RAW264.7 cells. Results In the in vitro culture of adult worms, the transcripts of CsMb increased with the increase of oxygen content. Oxidative stress conditions induced by H2O2 increased the levels of CsMb transcripts in a dose-dependent manner. Furthermore, CsMb catalyzed oxidation reactions in the presence of H2O2, and amino acid 34 of CsMb played an essential role in its reaction with H2O2. In addition, CsMb significantly reduced H2O2 and NO levels in LPS-activated macrophages, and CsMb downregulated iNOS and SOD expression in activated macrophages. Conclusion The present study is the first to investigate the peroxidase activity of CsMb. This investigation suggested that C. sinensis may decrease the redox activation of macrophages by CsMb expression to evade host immune responses. These studies contribute to a better understanding of the role of CsMb in the molecular mechanisms involved in ROS detoxification by C. sinensis.