Zhang Tong, Tsang Tom C, Harris David T
University of Arizona, Tucson, USA.
Biotechniques. 2002 Nov;33(5):1146-51. doi: 10.2144/02335dd09.
The long terminal repeat (LTR) of the human immunodeficiency virus (HIV) drives highly efficient gene expression in the presence of the transactivator, Tat. Thus, tat-containing vectors may be very useful tools in gene therapy. However information about the optimal way of delivering the tat gene is limited. In this study, we compared the effects of cis and trans expressions of the tat gene and its effects on HIV LTR-driven gene expression in different cell lines using non-viral vectors. The human interleukin-2 (IL-2) gene was used as a reporter gene under the control of the HIV2 LTR (pHIV2-IL-2). The tat gene, driven by a cytomegalovirus (CMV) promoter, was either co-transfected separately (pCMV-Tat) or inserted downstream of the IL-2 gene (pHIV2-IL-2-neo-C-Tat). Our results showed that HIV2 LTR-Tat-based vectors were much more potent than CMV promoter-based vectors in transient expression. The co-transfection of both plasmids was comparable to a single transfection of pHIV-IL-2-neo-C-Tat in both high and low transfection efficiency cells. In conclusion, the co-placement of HIV2 LTR and tat genes on a single plasmid allows for gene expression as efficiently as a two-plasmid system, suggesting that HIV2 LTR-Tat-based vectors may be attractive tools for gene therapy.
人类免疫缺陷病毒（HIV）的长末端重复序列（LTR）在反式激活因子Tat存在的情况下可驱动高效的基因表达。因此，含tat的载体可能是基因治疗中非常有用的工具。然而，关于递送tat基因的最佳方式的信息有限。在本研究中，我们使用非病毒载体比较了tat基因顺式和反式表达的效果及其对不同细胞系中HIV LTR驱动的基因表达的影响。人类白细胞介素-2（IL-2）基因在HIV2 LTR（pHIV2-IL-2）的控制下用作报告基因。由巨细胞病毒（CMV）启动子驱动的tat基因，要么单独共转染（pCMV-Tat），要么插入IL-2基因的下游（pHIV2-IL-2-neo-C-Tat）。我们的结果表明，基于HIV2 LTR-Tat的载体在瞬时表达方面比基于CMV启动子的载体更有效。在高转染效率细胞和低转染效率细胞中，两种质粒的共转染与单独转染pHIV-IL-2-neo-C-Tat相当。总之，HIV2 LTR和tat基因在单个质粒上的共同放置允许基因表达与双质粒系统一样高效，这表明基于HIV2 LTR-Tat的载体可能是基因治疗的有吸引力的工具。
