探討在肝癌細胞中CPAP 與HBx 蛋白的相互關係

Studying the relationship between CPAP and HBx in hepatocellular carcinoma

10.6844/NCKU.2013.01823

許魯信

肝細胞癌 ； CPAP ； B 型肝炎病毒X 蛋白 ； NF-kB ； HCC ； CPAP ； HBx ； NF-kB

成功大學藥理學研究所學位論文

2013年

碩士

洪良宜

英文

慢性發炎為導致肝癌的其中一個原因，而造成肝細胞慢性發炎的可能原因為肝炎病毒、酗酒或其它有毒的化學物質。持續活化的nuclear factor-kB (NF-kB)為參與肝臟細胞慢性發炎的重要因子。另一方面，於中心體功能上扮演重要角色的CPAP (Centrosomal protein P4.1-associated protein) 蛋白在先前的報導中被認為是NF-B 的轉錄共同活化者(transcriptional co-activator)。此外，過去文獻報導指出B 型肝炎病毒中的 X 蛋白 (HBx) 可增加NF-B 的活性。在本論文，我們發現GFP-CPAP 可協同性增加HBx 所調控的NF-kB 轉錄活性。利用siRNA抑制CPAP 表現，則減弱了此作用。利用免疫沉澱法(immunoprecipitation assay)顯示CPAP 與HBx 有交互作用。有趣的是，HBx 可透過CREB (cyclic adenosine monophosphate response element-binding protein) 來增加CPAP 的啟動子(promoter) 活性並造成CPAP 表現量的增加。此外，我們也發現在CPAP 大量表現的肝癌細胞株中，細胞增生能力有增加的情形。總結來說，我們的結果指出CPAP 可與HBx 合作來增加NF-kB 的活性，而這可能會導致肝癌的生成。

Hepatocellular carcinoma (HCC) is a type of tumor that results from chronic inflammation triggered by hepatotropic viruses, alcoholism or other toxic chemicals. Constitutive activation of nuclear factor-kB (NF-kB) is an important event involved in chronic inflammation of liver. CPAP (Centrosomal protein P4.1-associated protein), which plays an important role in centrosomal functions, is previously identified as a transcriptional co-activator of NF-kB. In addition, it was reported that the HBV X protein (HBx) of hepatitis B virus (HBV) can enhance the activation of NF-kB. In this study, we demonstrated that overexpression of GFP-CPAP synergistically increases the HBx-mediated NF-kB transcriptional activity, whereas knockdown expression of CPAP by siRNA attenuates this effect. Immunoprecipitation assay showed that CPAP interacts with HBx. Interestingly, HBx enhances the promoter activity of CPAP may be through the interaction with cyclic adenosine monophosphate response element-binding protein (CREB). Stably expression of CPAP in HCC cell lines increased the cell proliferation ability. Taken together, our results indicate that CPAP can cooperate with HBx to increase the NF-kB activity, which may contribute to hepatocarcinogenesis.

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