A newly modified SCG10 promoter and Cre/loxP-mediated gene amplification system achieve highly specific neuronal expression in animal brains.
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.
Namikawa
K
K
Department of Anatomy and Neurobiology, Osaka City University, Graduate School of Medicine, Abenoku, Osaka, Japan.
Murakami
K
K
Okamoto
T
T
Okado
H
H
Kiyama
H
H
D016475:3T3 Cells; D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001253:Astrocytes / Q000378:metabolism; D001921:Brain / Q000378:metabolism*; D018920:Coculture Techniques; D004742:Enhancer Elements, Genetic; D005784:Gene Amplification; D005818:Genetic Engineering; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D019426:Integrases / Q000235:genetics*; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D051381:Rats; D037941:Silencer Elements, Transcriptional / Q000235:genetics; D050777:Stathmin; D014161:Transduction, Genetic
D008565:Membrane Proteins; C063314:STMN2 protein, human; D050777:Stathmin; D049452:Green Fluorescent Proteins; C045073:Cre recombinase; D019426:Integrases