A Baculovirus Superinfection System: Efficient Vehicle for Gene Transfer into Drosophila S2 Cells

The baculovirus expression vector system is considered to be a safe, powerful, but cell-lytic heterologous protein expression system in insect cells. We show here that there is a new baculovirus system for efficient gene transfer and expression using the popular and genetically well-understood Drosophila S2 cells. The recombinant baculovirus was constructed to carry an enhanced green fluorescent protein under the control of polyhedrin promoter as a fluorescent selection marker in the Sf21 cell line. Recombinant baculoviruses were then used to transduce S2 cells with target gene expression cassettes containing a Drosophila heat shock protein 70, an actin 5C, or a metallothionein promoter. Nearly 100% of the S2 cells showed evidence of gene expression after infection. The time course for the optimal protein expression peaked at 24 to 36 h postinfection, which is significantly earlier than a polyhedrin-driven protein expression in Sf21 cells. Importantly, S2 cells did not appear to be lysed after infection, and the protein expression levels are comparable to those of proteins under the control of polyhedrin promoter in several lepidopteran cell lines. Most surprisingly, S2 cells permit repetitive infections of multiple baculoviruses over time. These findings clearly suggest that this baculovirus-S2 system may effect the efficient gene transfer and expression system of the well-characterized Drosophila S2 cells.

杆状病毒表达载体系统（baculovirus expression vector system）被公认为一类安全、高效但具备细胞裂解特性的昆虫细胞异源蛋白表达系统。本研究表明，可借助应用广泛且遗传背景明晰的果蝇S2细胞（Drosophila S2 cells），构建一套新型高效的基因转移与蛋白表达杆状病毒系统。研究人员构建了携带受多角体蛋白启动子（polyhedrin promoter）调控的增强型绿色荧光蛋白（enhanced green fluorescent protein）的重组杆状病毒，将其作为Sf21细胞系（Sf21 cell line）中的荧光筛选标记。随后利用该重组杆状病毒转导S2细胞，所搭载的靶基因表达盒（target gene expression cassettes）分别受果蝇热休克蛋白70（Drosophila heat shock protein 70）、肌动蛋白5C（actin 5C）或金属硫蛋白启动子（metallothionein promoter）调控。感染后，几乎100%的S2细胞均可检测到基因表达现象。最优蛋白表达的时间进程于感染后24至36小时达到峰值，这比Sf21细胞中多角体蛋白驱动的蛋白表达时间显著提前。尤为关键的是，S2细胞在感染后并未出现裂解情况，且蛋白表达水平可与多种鳞翅目细胞系（lepidopteran cell lines）中多角体蛋白启动子调控下的蛋白表达水平相媲美。最令人意外的是，S2细胞可在培养周期内重复接受多种杆状病毒的感染。上述研究结果充分证实，该杆状病毒-S2系统可作为一套适配遗传特征明确的果蝇S2细胞的高效基因转移与蛋白表达系统。