Namikawa K, Murakami K, Okamoto T, Okado H, Kiyama H
Department of Anatomy and Neurobiology, Osaka City University, Graduate School of Medicine, Abenoku, Osaka, Japan.
Gene Ther. 2006 Aug;13(16):1244-50. doi: 10.1038/sj.gt.3302779. Epub 2006 Apr 20.
We designed a new promoter that drives transgene expression in an exclusively neuron-specific manner. The promoter of superior cervical ganglion10 (SCG10), expressed in neurons, was further modified to enhance its neuron specificity and activity by changing its length and fusing a multiple neuronal restrictive silencer element (NRSE) to its upstream or downstream regions. The promoter, which contained 2 kb original promoter length and two extra NRSEs in its downstream region, eventually exhibited remarkable neuron specificity as well as strong activity. To further amplify the promoter activity, the promoter was introduced into a Cre recombinase (Cre)-expressing adenovirus, and subsequent combination with Cre-inducible enhanced green fluorescence protein (EGFP)-expressing adenovirus vector, which has much stronger general promoter, resulted in a remarkably strong gene expression exclusively in neuronal cells of mixed cultures and in an animal model. This system is also applicable to astrocyte-specific expression; for instance, by changing the Cre promoter cassette to an astrocyte-specific promoter. The present relatively compact promoter combined with Cre/loxP system could be useful for a wide range of transgene experiments in vivo as well as for clinical applications.
我们设计了一种新型启动子，它能以完全神经元特异性的方式驱动转基因表达。在神经元中表达的颈上神经节10（SCG10）启动子，通过改变其长度并在其上游或下游区域融合多个神经元限制性沉默元件（NRSE），进一步进行修饰以增强其神经元特异性和活性。该启动子在其下游区域包含2 kb的原始启动子长度和两个额外的NRSE，最终表现出显著的神经元特异性以及强大的活性。为了进一步增强启动子活性，将该启动子导入表达Cre重组酶（Cre）的腺病毒中，随后与具有更强通用启动子的Cre诱导型增强绿色荧光蛋白（EGFP）表达腺病毒载体相结合，导致在混合培养物的神经元细胞以及动物模型中仅产生非常强的基因表达。该系统也适用于星形胶质细胞特异性表达；例如，通过将Cre启动子盒更换为星形胶质细胞特异性启动子。目前这种相对紧凑的启动子与Cre/loxP系统相结合，可用于体内广泛的转基因实验以及临床应用。
