Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, Institute of Medical Microbiology and Institutes of Biomedical Sciences, Shanghai Medical College of Fudan University, Shanghai, China.
Department of Chemistry, Institutes of Biomedical Sciences, Fudan University, Shanghai, China.
Proteome Sci. 2010 Jun 7;8:28. doi: 10.1186/1477-5956-8-28.
Hepatitis B virus (HBV) is a major cause of liver infection in human. Because of the lack of an appropriate cell culture system for supporting HBV infection efficiently, the cellular and molecular mechanisms of hepadnavirus infection remain incompletely understood. Duck heptatitis B virus (DHBV) can naturally infect primary duck hepatocytes (PDHs) that provide valuable model systems for studying hepadnavirus infection in vitro. In this report, we explored global changes in cellular protein expression in DHBV infected PDHs by two-dimension gel electrophoresis (2-DE) combined with MALDI-TOF/TOF tandem mass spectrometry (MS/MS).
The effects of hepadnavirus infection on hepatocytes were investigated in DHBV infected PDHs by the 2-DE analysis. Proteomic profile of PDHs infected with DHBV were analyzed at 24, 72 and 120 h post-infection by comparing with uninfected PDHs, and 75 differentially expressed protein spots were revealed by 2-DE analysis. Among the selected protein spots, 51 spots were identified corresponding to 42 proteins by MS/MS analysis; most of them were matched to orthologous proteins of Gallus gallus, Anas platyrhynchos or other avian species, including alpha-enolase, lamin A, aconitase 2, cofilin-2 and annexin A2, etc. The down-regulated expression of beta-actin and annexin A2 was confirmed by Western blot analysis, and potential roles of some differentially expressed proteins in the virus-infected cells have been discussed.
Differentially expressed proteins of DHBV infected PDHs revealed by 2-DE, are involved in carbohydrate metabolism, amino acid metabolism, stress responses and cytoskeleton processes etc, providing the insight to understanding of interactions between hepadnavirus and hepatocytes and molecular mechanisms of hepadnavirus pathogenesis.
乙型肝炎病毒（HBV）是人类肝脏感染的主要原因。由于缺乏有效的细胞培养系统来支持 HBV 的高效感染，因此对嗜肝病毒感染的细胞和分子机制仍不完全了解。鸭乙型肝炎病毒（DHBV）可自然感染原代鸭肝细胞（PDH），为研究体外嗜肝病毒感染提供了有价值的模型系统。在本报告中，我们通过二维凝胶电泳（2-DE）结合 MALDI-TOF/TOF 串联质谱（MS/MS）研究了 DHBV 感染 PDH 后细胞蛋白表达的全局变化。
通过 2-DE 分析研究了 DHBV 感染 PDH 对肝细胞的影响。通过与未感染 PDH 相比，在感染后 24、72 和 120 小时分析 DHBV 感染 PDH 的蛋白质组谱，通过 2-DE 分析显示 75 个差异表达的蛋白斑点。在选择的蛋白斑点中，通过 MS/MS 分析鉴定了 51 个与 42 个蛋白相对应的蛋白斑点；其中大多数与 Gallus gallus、Anas platyrhynchos 或其他禽类的同源蛋白相匹配，包括α-烯醇酶、核纤层蛋白 A、 aconitase 2、cofilin-2 和 annexin A2 等。Western blot 分析证实了β-肌动蛋白和 annexin A2 的下调表达，并讨论了一些差异表达蛋白在病毒感染细胞中的潜在作用。
2-DE 显示的 DHBV 感染 PDH 的差异表达蛋白涉及碳水化合物代谢、氨基酸代谢、应激反应和细胞骨架过程等，为理解嗜肝病毒与肝细胞的相互作用和嗜肝病毒发病机制的分子机制提供了深入的了解。
