Santi Nina, Sandtrø Ane, Sindre Hilde, Song Haichen, Hong Jiann-Ruey, Thu Beate, Wu Jen-Leih, Vakharia Vikram N, Evensen Øystein
Section for Pathology, National Veterinary Institute, 0033 Oslo, Norway.
Virology. 2005 Nov 10;342(1):13-25. doi: 10.1016/j.virol.2005.07.028. Epub 2005 Aug 26.
Infectious pancreatic necrosis virus (IPNV), the causative agent of a highly infectious disease in salmonid fish, encodes a small non-structural protein designated VP5. This protein contains Bcl-2 homologous domains and inhibits apoptosis when expressed in cell culture. We have previously reported the generation of three VP5 mutants of IPNV-Sp serotype, using reverse genetics (Santi, N., Song, H., Vakharia, V.N., Evensen, Ø., 2005. Infectious pancreatic necrosis virus VP5 is dispensable for virulence and persistence. J. Virol. 79 (14), 9206-9216). The wild-type rNVI15 virus encodes a truncated 12-kDa VP5 protein, rNVI15-15K encodes a full-length 15-kDa VP5, whereas rNVI15-DeltaVP5 is deficient in VP5 expression. In the present report, the role of VP5 in apoptosis was assessed both in vitro and in vivo, using the recombinant IPNV strains. Apoptosis was observed in hepatocytes of Atlantic salmon post-smolts challenged with all three VP5 mutant viruses. Using a double-labeling technique to detect apoptotic cells and IPNV antigens, we found that viral antigen and apoptotic cells co-distributed. In addition, numerous double-positive cells were seen. The recombinant viruses also induced apoptosis in infected cell cultures, and the morphology and membrane integrity of infected cells at different time points was similar. In summary, these results indicate that IPNV induces apoptosis in infected cell cultures and in fish, independent of VP5 expression. However, substitutions of putative functionally important amino acids in the BH2 domain of VP5 of IPNV-Sp strains were identified, which might influence the anti-apoptosis effect of the protein, and partly explain the apparent absence of this specific function.
传染性胰腺坏死病毒（IPNV）是鲑科鱼类一种高度传染性疾病的病原体，它编码一种名为VP5的小非结构蛋白。该蛋白含有Bcl-2同源结构域，在细胞培养中表达时可抑制细胞凋亡。我们之前曾报道过利用反向遗传学方法构建了IPNV-Sp血清型的三种VP5突变体（桑蒂，N.，宋，H.，瓦卡里亚，V.N.，埃文森，Ø.，2005年。传染性胰腺坏死病毒VP5对毒力和持续性并非必需。《病毒学杂志》79(14)，9206 - 9216）。野生型rNVI15病毒编码一个截短的12 kDa VP5蛋白，rNVI15 - 15K编码全长15 kDa的VP5，而rNVI15 - ΔVP5缺乏VP5表达。在本报告中，使用重组IPNV毒株在体外和体内评估了VP5在细胞凋亡中的作用。在用所有三种VP5突变病毒攻击的大西洋鲑后幼鱼的肝细胞中观察到了细胞凋亡。使用双标记技术检测凋亡细胞和IPNV抗原，我们发现病毒抗原和凋亡细胞共分布。此外，还观察到大量双阳性细胞。重组病毒在感染的细胞培养物中也诱导了细胞凋亡，并且在不同时间点感染细胞的形态和膜完整性相似。总之，这些结果表明IPNV在感染的细胞培养物和鱼类中诱导细胞凋亡，与VP5表达无关。然而，已鉴定出IPNV - Sp毒株VP5的BH2结构域中假定功能重要氨基酸的替代，这可能会影响该蛋白的抗凋亡作用，并部分解释了这种特定功能明显缺失的原因。
