Department of Oncology, Tangdu Hospital, The Fourth Military Medical University, Xi'an, People's Republic of China.
J Med Virol. 2010 May;82(6):920-8. doi: 10.1002/jmv.21753.
Hepatitis B virus (HBV) infection is closely correlated with the development of hepatocellular carcinoma (HCC), in which hepatitis B virus X protein (HBx) plays crucial roles. HBx is believed to be a multifunctional oncoprotein. It has been reported that the calpain small subunit 1 (Capn4) is upregulated in the HCC tissues and involved in the metastasis of HCC. Therefore, we suppose that HBx may promote hepatoma cell migration through Capn4. In the present study, we investigated the effect of HBx on regulating Capn4 expression in human HCC cells. Our data showed that HBx could increase promoter activity of Capn4 and upregulate the expression of Capn4 at the levels of mRNA and protein in human hepatoma HepG2 (or H7402) cells using luciferase reporter gene assay, real-time quantitative RT-PCR assay and Western blot analysis. While, the RNA interference targeting HBx mRNA was able to abolish the upregulation. Interestingly, we found that the inhibition of nuclear factor-kappaB (NF-kappaB) mediated by siRNA targeting NF-kappaB/p65 mRNA or PDTC (an inhibitor of NF-kappaB) could attenuate the upregulation of Capn4. While, HBx failed to increase the promoter activity of Capn4 in hepatoma cells when the putative NF-kappaB binding site of the Capn4 promoter was mutant, suggesting that NF-kappaB is involved in the activation of Capn4 mediated by HBx. In function, wound healing assay showed that HBx could significantly enhance the migration ability of HepG2 cells through upregulating Capn4. Thus, we conclude that HBx upregulate Capn4 through NF-kappaB/p65 to promote migration of hepatoma cells.
乙型肝炎病毒（HBV）感染与肝细胞癌（HCC）的发展密切相关，其中乙型肝炎病毒 X 蛋白（HBx）起着关键作用。HBx 被认为是一种多功能癌蛋白。据报道，钙蛋白酶小亚基 1（Capn4）在 HCC 组织中上调，并参与 HCC 的转移。因此，我们假设 HBx 可能通过 Capn4 促进肝癌细胞迁移。在本研究中，我们研究了 HBx 对调节人 HCC 细胞中 Capn4 表达的影响。我们的数据表明，HBx 可以通过荧光素酶报告基因检测、实时定量 RT-PCR 检测和 Western blot 分析，增加 Capn4 启动子活性，并上调人肝癌 HepG2（或 H7402）细胞中 Capn4 的 mRNA 和蛋白水平。而靶向 HBx mRNA 的 RNA 干扰能够消除这种上调。有趣的是，我们发现靶向 NF-kappaB/p65 mRNA 的 siRNA 或 PDTC（NF-kappaB 的抑制剂）抑制核因子-kappaB（NF-kappaB）可以减弱 Capn4 的上调。而当 Capn4 启动子的假定 NF-kappaB 结合位点发生突变时，HBx 未能增加肝癌细胞中 Capn4 的启动子活性，这表明 NF-kappaB 参与了 HBx 介导的 Capn4 激活。在功能上，划痕愈合实验表明，HBx 通过上调 Capn4 显著增强 HepG2 细胞的迁移能力。因此，我们得出结论，HBx 通过 NF-kappaB/p65 上调 Capn4 促进肝癌细胞迁移。
