Turner P C, Watkins P A, Zaitlin M, Wilson T M
Department of Biochemistry, University of Liverpool, P.O. Box 147, Liverpool L69 3BX, United Kingdom.
Virology. 1987 Oct;160(2):515-7. doi: 10.1016/0042-6822(87)90029-8.
Xenopus laevis oocytes were microinjected with tobacco mosaic virus (TMV) particles or unencapsidated TMV RNA at equivalent RNA concentrations. After a 5-hr incubation, batches of 10 oocytes from each treatment were frozen and lysed, and the supernatants were subjected to SDS-PAGE and Western immunoblotting with antiserum raised against a TMV RNA-encoded polypeptide of mol wt 126,000 (126K), purified from TMV-infected tobacco. Cross-reaction with a polypeptide of 126K synthesized in vitro, in a rabbit reticulocyte cell-free system programmed with TMV RNA, confirmed the specificity of the antiserum. Oocytes microinjected with TMV particles produced at least as much immunoreactive 126K protein as those injected with TMV RNA. We conclude that intact virus particles were disassembled, at least in part, to release some functional RNA within the cytoplasm of these animal cells. Hypotheses which postulate a specific role for cellulose cell walls, or the external surface of the plasmalemma, in virus disassembly seem unable to explain this observation. We favor an intracellular site and a more universal mechanism for ribonucleocapsid disassembly. This result may also affect our views on factors which determine the host range of plant viruses.
将烟草花叶病毒(TMV)颗粒或未包裹的TMV RNA以相等的RNA浓度显微注射到非洲爪蟾卵母细胞中。孵育5小时后,将每种处理的10个卵母细胞一批进行冷冻和裂解,然后将上清液进行SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)和用针对从感染TMV的烟草中纯化的分子量为126,000(126K)的TMV RNA编码多肽产生的抗血清进行Western免疫印迹分析。用在含有TMV RNA编程的兔网织红细胞无细胞系统中体外合成的126K多肽进行交叉反应,证实了抗血清的特异性。显微注射TMV颗粒的卵母细胞产生的免疫反应性126K蛋白至少与注射TMV RNA的卵母细胞一样多。我们得出结论,完整的病毒颗粒至少部分地被拆解,以在这些动物细胞的细胞质中释放一些功能性RNA。假设纤维素细胞壁或质膜外表面在病毒拆解中起特定作用的假说似乎无法解释这一观察结果。我们倾向于一种细胞内位点和一种更普遍的核糖核蛋白体拆解机制。这一结果也可能会影响我们对决定植物病毒宿主范围的因素的看法。