Quantitative Proteomic Analysis of Exosome Protein Content Changes Induced by Hepatitis B Virus in Huh-7 Cells Using SILAC Labeling and LC鈥揗S/MS

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• 作者：Xue Zhao ; Yanxin Wu ; Jinlin Duan ; Yanchun Ma ; Zhongliang Shen ; Lili Wei ; Xiaoxian Cui ; Junqi Zhang ; Youhua Xie ; Jing Liu
• 刊名：Journal of Proteome Research
• 出版年：2014
• 出版时间：December 5, 2014
• 年：2014
• 卷：13
• 期：12
• 页码：5391-5402
• 全文大小：466K
• ISSN：1535-3907

文摘

Hepatitis B virus (HBV) infection could cause hepatitis, liver cirrhosis, and hepatocellular carcinoma. HBV-mediated pathogenesis is only partially understood, but X protein (HBx) reportedly possesses oncogenic potential. Exosomes are small membrane vesicles with diverse functions released by various cells including hepatocytes, and HBV harnesses cellular exosome biogenesis and export machineries for virion morphogenesis and secretion. Therefore, HBV infection might cause changes in exosome contents with functional implications for both virus and host. In this work, exosome protein content changes induced by HBV and HBx were quantitatively analyzed by SILAC/LC鈥揗S/MS. Exosomes prepared from SILAC-labeled hepatoma cell line Huh-7 transfected with HBx, wildtype, or HBx-null HBV replicon plasmids were analyzed by LC鈥揗S/MS. Systematic analyses of MS data and confirmatory immunoblotting showed that HBx overexpression and HBV, with or without HBx, replication in Huh-7 cells indeed caused marked and specific changes in exosome protein contents. Furthermore, specific changes in protein contents were also detected in exosomes purified from HBV-infected patients鈥?sera compared with control sera negative for HBV markers. These results illustrate a new aspect of interactions between HBV and the host and provide the foundation for future research into roles played by exosomes in HBV infection and pathogenesis.

Keywords:

HBV; HBx; exosome; hepatocyte; SILAC; patient serum; VCP