核衣壳蛋白VP39融合TAT转导肽对杆状病毒转导哺乳动物细胞的影响

Tat transduction peptide displayed on the nucleocapsid improved the baculovirus transduction of mammalian cells

摘要:

为了提高昆虫杆状病毒在哺乳动物细胞中转导基因的效率, 构建了重组杆状病毒AcRed-tat和AcRed。两者都能在哺乳动物细胞内表达红色荧光蛋白作为报告基因。同时, AcRed-tat带有HIV-1 Tat转导肽、病毒主要衣壳蛋白基因vp39及增强型绿色荧光蛋白(egfp)三者的融合基因, 并由杆状病毒多角体启动子表达, 能够在昆虫细胞中表达该Tat融合蛋白, 并掺入子代病毒粒子。而AcRed作为相应的对照病毒, 带有多角体启动子表达vp39和egfp的融合基因。2株病毒分别转导哺乳动物细胞后, 利用流式细胞仪检测报告基因的表达水平, 发现在CHO和Vero细胞中AcRed-Tat介导的报告基因表达水平明显高于AcRed, 而在HEK293细胞中2株病毒介导的报告基因表达水平差异不显著。结果表明Tat转导肽可以提高杆状病毒对一部分哺乳动物细胞的转导效率, 为改进杆状病毒-哺乳动物细胞转导载体提供了新的思路。

Abstract:

In order to improve the transduction efficiency of insect baculovirus in mammalian cells, we constructed two recombinant baculoviruses, AcRed-tat and AcRed. Both viruses expressed red fluorescence protein gene (dsRed) as a reporter in mammalian cell lines. AcRed-tat also contained the coding sequence of HIV-1 Tat transduction peptide fused with viral major capsid protein gene vp39 and enhanced green fluorescence gene (egfp) driven by virus polyhedrin promoter. It expressed the Tat fusion protein in infected insect cells, which was incorporated into the nucleocapsids of progeny virus. As a control, AcRed had the fusion gene of vp39 and egfp driven by polyhedrin promoter. Flow cytometry analysis demonstrated that although similar level of red fluorescence was produced in HEK23 cells transduced by the two recombinant viruses, significantly higher red fluorescence level was seen in CHO and Vero cells transduced by AcRed-tat than that by AcRed. These results suggested that Tat transduction peptide might improve the baculovirus-mediated gene expression in some mammalian cells. Our work provided a new approach to improve baculovirus as a gene delivery vector for mammalian cells.