Study for mechanisms of NF-κB activation by hepatitis C viral core protein

慢性丙型肝炎病毒（HCV）感染被认为会导致慢性肝炎、肝硬化和肝细胞癌的发展。因此，我们试图揭示NF-κB被HCV核心蛋白激活的分子机制，该核心蛋白被认为与该病毒的持续感染有关。利用酵母双杂交筛选系统，我们获得了几个候选的与核心相互作用的细胞因子。其中一个候选基因是Sp110b，据报道它是视黄酸受体（RAR）转录辅因子Sp110的剪接变体。由于我们观察到核心增强了RAR的转录活性，因此对核心与Sp110b之间的关系进行了更细致的分析。Sp110b mRNA的表达量高于Sp110。我们还观察到，过量产生Sp110b抑制了rar依赖的转录激活，这表明Sp110b是rar依赖转录的负辅因子。我们发现，原位于细胞核的Sp110b亚细胞定位在核周细胞质区，当核在细胞内共产生时。这种变化以及rar依赖性转录激活的增强通过与核心结合所需的Sp110b最小区域的共同产生而恢复，这表明核心和Sp110b之间的相互作用对这些现象很重要。由于一个转录辅助因子已被发现与多个转录事件相关，我们现在研究Sp110b对NF-κB活化的贡献。此外，我们建立了HCV基因组复制子的细胞系，包括整个HCV基因组的复制子被复制和维持，以检验核心在这些细胞中的作用。利用该细胞系分析了Sp110b在HCV基因组复制中的功能。

Chronic infection of hepatitis C virus (HCV) is considered to cause the development of chronic hepatitis, liver cirrohsis, and hepatocellular carcinoma. We tried, therefore, to reveal the molecular mechanism of NF-κB activation by the HCV core protein (the core) which has been suggested to be related with the persistent infection of this virus. Using the yeast two hybrid screening system, we obtained several candidates of cellular factors interacting with the core. One of the candidates was Sp110b, which was reported to be a splicing variant of Sp110, a transcriptional cofactor of retinoic acid receptor (RAR). As we observed that the core enhanced the transcriptional activity of RAR, finer analyses for the relationship between the core and Sp110b were performed. Expression level of Sp110b mRNA was higher than that of Sp110. We also observed that overproduction of Sp110b suppressed the RAR-dependent transcriptional activation, suggesting that Sp110b is a negative cofactor of RAR-dependent transcription. We found that the subcellular localization of Sp110b which was originally located in the nucleus was changed to the perinuclear cytoplasmic region, when the core was coproduced in the cells. That change as well as the enhancement of RAR-dependent transcriptional activation was reverted by the coproduction of minimum region of Sp110b required for binding with the core, suggesting that the interaction between the core and Sp110b is important for those phenomenons. As a transcriptional cofactor has been revealed to be related with several transcriptional events, we now examine the contribution of Sp110b to the activation of NF-κB by the core.Furthermore, we have established the cell line in which HCV genomic replicon including whole HCV genome was replicated and maintained in order to examine the effect of the core in such cells. The analysis of function of Sp110b on the HCV genome replication is underway by using this cell line.