神經壞死病毒(Nervous Necrosis Virus, NNV)屬於野田病毒(Betanodavirus)的一種，可感染多種高經濟性魚種如石斑魚。在幼苗期具有高致死率，對水產養殖業造成嚴重的經濟損失。先前研究中RGNNV感染石斑魚細胞株會造成粒線體膜電位消失及細胞壞死，並且病毒感染也伴隨ROS (reactive oxygen species)產生。此外，B1在病毒感染後可以降低病毒蛋白表現量及減緩細胞死亡，但目前B1是否能夠調節ROS及如何延遲細胞死亡，仍然不明。本篇論文發現表現B1能誘發少量O2-的產生，同時微量的氧化壓力會誘發抗氧化酵素Cu/Zn SOD、Mn SOD以及catalase的表現量上升及細胞周期停滯於G1期。另一方面，B1能減少H2O2、H2O2誘導劑tBHQ以及凋亡誘導劑Act D (Actinomycin D) 所產生的細胞凋亡。此外，B1進核與否會減弱抵抗H2O2的能力，所以B1進核或許對B1的功能是一個重要的影響關鍵。最後藉由微矩陣晶片分析表現B1後轉錄體的差異，希望能找出有關ROS以及抵抗細胞死亡的線索。根據實驗結果，得知B1可能是一種具有多種功能的蛋白，除了調節氧化壓力外也會造成細胞周期停滯並抑制細胞凋亡。這些發現可以在未來對野田病毒的感染、致病機制有更深的理解與應用。

As we known, some fish species are infected by betanodavirus such as grouper, which results in severe mortality and significant economic losses on the aquaculture industry. In previous studies, RGNNV can induce necrotic cell death via mitochondrial membrane (MMP) loss in grouper cells. The RGNNV B1 is belonging to early expression gene which plays a crucial role on anti-apoptotic function and reduces viral replication. But, whether B1 overexpression can regulate ROS is still unknown. And how to reduce cell death with virus infection is also unclear. In this study, we found that the B1 can induce superoxide anion (O2- ) production at 12h. And then the B1 can up-regulate some anti-oxidant enzymes such as Cu/Zn SOD, Mn SOD, catalase and arrest the cell cycle via oxidative stress response. On the other hand, the B1 can reduce cell death with treat H2O2, tBHQ (H2O2 inducer) and Act D (Actinomycin D), an apoptosis inducer. Furthermore, we still don’t know whether B1 protein targeting into nucleus is correlative to anti-death function. In the result, we found that B1 targeting into nucleus can correlate to B1 function. Finally, we analyzed the transcription level by cDNA microarray after EYFP-B1 transfection and try to find some hint for ROS and anti-cell death studies. In the result, the B1 protein may play a multifunction roles, which not only arrest cell cycle but also induce ROS production and cell death. This first finding may provide an insight into the molecular pathogenesis of betanodavirus infection in future.
Keywords: Betanodavirus, nonstructural protein B1, reactive oxygen species (ROS), oxidative stress respone, anti-oxidant enzyme, cell cycle arrest, anti-apoptosis.

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