Rodriguez J F, Smith G L
Sir William Dunn School of Pathology, University of Oxford, UK.
Nucleic Acids Res. 1990 Sep 25;18(18):5347-51. doi: 10.1093/nar/18.18.5347.
A novel method has been developed to study the functional roles of individual vaccinia virus gene products that is neither limited by the possible essentiality of the target gene nor by the availability of conditional lethal mutants. The system utilises the E. coli lac repressor protein, the operator sequence to which it binds and the specific inducer IPTG. It allows the generation of recombinant viruses in which the expression of any chosen gene, and hence virus replication, can be externally controlled. In principle, this system is broadly applicable to the functional analysis of genes in any large DNA virus. This approach has demonstrated that the gene encoding the 14 kDa membrane protein of vaccinia virus is non-essential for the production of infectious intracellular virus particles, but essential for the envelopment of intracellular virions by Golgi membrane and for egress of mature extracellular viral particles. This is the first vaccinia virus protein shown to be specifically required for these processes. In vivo this system may prove useful as a means of attenuating recombinant vaccinia virus vaccines by preventing virus spread without reducing the amount of the foreign antigen expressed in each infected cell. Attenuation of other live virus vaccines may be developed in a similar way.
已开发出一种新方法来研究痘苗病毒单个基因产物的功能作用，该方法既不受靶基因可能的必需性限制，也不受条件致死突变体可用性的限制。该系统利用大肠杆菌乳糖阻遏蛋白、其结合的操纵序列和特异性诱导剂IPTG。它允许产生重组病毒，其中任何选定基因的表达以及病毒复制都可以在外部进行控制。原则上，该系统广泛适用于任何大型DNA病毒中基因的功能分析。这种方法已经证明，编码痘苗病毒14 kDa膜蛋白的基因对于感染性细胞内病毒颗粒的产生不是必需的，但对于高尔基体膜包裹细胞内病毒粒子以及成熟细胞外病毒粒子的释放是必需的。这是第一个被证明在这些过程中具有特异性需求的痘苗病毒蛋白。在体内，该系统可能被证明是一种有用的方法，可通过防止病毒传播而不减少每个感染细胞中表达的外源抗原量来减毒重组痘苗病毒疫苗。其他活病毒疫苗的减毒也可以通过类似的方式进行开发。
