Department of Microbiology, Molecular Genetics and Immunology, Mail Stop 3029, 3901 Rainbow Blvd., Kansas City, KS 66160, USA.
J Virol. 2010 Mar;84(6):2687-96. doi: 10.1128/JVI.01917-09. Epub 2009 Dec 30.
Aleutian mink disease virus (AMDV) is currently the only known member of the genus Amdovirus in the family Parvoviridae. It is the etiological agent of Aleutian disease of mink. We have previously shown that a small protein with a molecular mass of approximately 26 kDa was present during AMDV infection and following transfection of capsid expression constructs (J. Qiu, F. Cheng, L. R. Burger, and D. Pintel, J. Virol. 80:654-662, 2006). In this study, we report that the capsid proteins were specifically cleaved at aspartic acid residue 420 (D420) during virus infection, resulting in the previously observed cleavage product. Mutation of a single amino acid residue at D420 abolished the specific cleavage. Expression of the capsid proteins alone in Crandell feline kidney (CrFK) cells reproduced the cleavage of the capsid proteins in virus infection. More importantly, capsid protein expression alone induced active caspases, of which caspase-10 was the most active. Active caspases, in turn, cleaved capsid proteins in vivo. Our results also showed that active caspase-7 specifically cleaved capsid proteins at D420 in vitro. These results suggest that viral capsid proteins alone induce caspase activation, resulting in cleavage of capsid proteins. We also provide evidence that AMDV mutants resistant to caspase-mediated capsid cleavage increased virus production approximately 3- to 5-fold in CrFK cells compared to that produced from the parent virus AMDV-G at 37 degrees C but not at 31.8 degrees C. Collectively, our results indicate that caspase activity plays multiple roles in AMDV infection and that cleavage of the capsid proteins might have a role in regulating persistent infection of AMDV.
阿留申病病毒(AMDV)是细小病毒科细小病毒属中目前唯一已知的成员。它是引起水貂阿留申病的病原体。我们之前已经表明,在 AMDV 感染和衣壳表达构建体转染后,存在一种分子量约为 26 kDa 的小蛋白(J. Qiu、F. Cheng、L. R. Burger 和 D. Pintel,J. Virol. 80:654-662, 2006)。在这项研究中,我们报告说,衣壳蛋白在病毒感染过程中在天冬氨酸残基 420 处(D420)被特异性切割,导致先前观察到的切割产物。在 D420 处单个氨基酸残基的突变消除了特异性切割。单独在猫肾细胞(CrFK)中表达衣壳蛋白复制了病毒感染中衣壳蛋白的切割。更重要的是,单独表达衣壳蛋白诱导活性半胱天冬酶,其中半胱天冬酶-10 最活跃。活性半胱天冬酶反过来在体内切割衣壳蛋白。我们的结果还表明,活性半胱天冬酶-7 在体外特异性地在 D420 处切割衣壳蛋白。这些结果表明,单独的病毒衣壳蛋白诱导半胱天冬酶激活,导致衣壳蛋白的切割。我们还提供了证据表明,与亲本病毒 AMDV-G 相比,对 caspase 介导的衣壳切割具有抗性的 AMDV 突变体在 37°C 下但不在 31.8°C 下在 CrFK 细胞中产生的病毒产量增加了约 3-5 倍。总的来说,我们的结果表明,半胱天冬酶活性在 AMDV 感染中发挥多种作用,并且衣壳蛋白的切割可能在调节 AMDV 的持续性感染中发挥作用。