Molecular analysis on the tumorigenicity of hepatitis C virus nonstructural protein NS3

为了检测丙型肝炎病毒NS3对细胞的影响，我们采用脂质体转染法。含编码丙型肝炎病毒NS3基因5‘端半序列的转基因载体(丙型肝炎病毒NS3-N)在裸鼠体内迅速增殖并形成肿瘤。但与抑癌基因p53共转染后，未见致瘤性。结果提示，丙型肝炎病毒NS3对P53的功能有一定影响。为了研究丙型肝炎病毒NS3与P53的相互作用，我们制备了在细菌中表达的融合蛋白GST-丙型肝炎病毒-NS3。在细胞提取物的体外结合实验中，我们没有检测到NS3与P53的结合。这不排除丙型肝炎病毒NS3和P53之间弱的或间接的相互作用的可能性。实际上，我们发现了能与丙型肝炎病毒NS3结合的90 kDa蛋白。另一方面，我们试图利用丙型肝炎病毒相关病毒的复制系统寻找有效的抗病毒药物，即乙型脑炎病毒(JEV)。一种被测试的试剂，呋喃萘醌(FNQ)显示出抗病毒活性(Takegami等人，1998年)。为了延缓丙型肝炎病毒的致瘤，抑制病毒复制和病毒与宿主细胞之间的某些反应是很重要的。进一步分析丙型肝炎病毒NS3与包括P53在内的宿主蛋白的相互作用，以及FNQ的抗病毒作用机制是十分必要的。

In order to examine the influence of hepatitis C virus (HCV) NS3 to the cells, we used transfection method using lipofectin. Transfectant containing 5'half sequence of the HCV genome segment encoding NS3 (HCV-NS3-N) rapidly proliferated and formed tumors in nude mice. However, after the cotransfection with tumor suppressor gene p53 transfectants (HCV-NS3-N+p53) did not show any tumorigenicity. The result suggest that HCV-NS3 has some influence to the function of p53. To investigate the interaction between HCV-NS3 and p53, we prepared the fusion-protein GST-HCV-NS3 expressed in the bacteria. In the binding experiment in vitro using cell extracts, we did not detect the binding of NS3 to p53. This does not exclude the possibility of the weak or indirect interaction between HCV-NS3 and p53. Actually we found 90 kDa protein which could bind to HCV-NS3. On the other hand, we tried to find the effective antiviral drug using the replication system of HCV-related virus, i.e. Japanese encephalitis virus (JEV). One of tested reagents, furanonaphtoquinone (FNQ) showed antiviral activity (Takegami et al., 1998). To retard the tumorigenesis by HCV,it is important to inhibit viral replication and some reactions between virus and host cells. Further analyzes on the interaction of HCV-NS3 and host proteins including p53, and mechanism of antiviral effect of FNQ are essential.