Synthetic promoter for efficient and muscle-specific expression of exogenous genes

Abstract

Synthetic promoters (SPs) have many advantages over their natural counterparts, especially with regard to transcriptional activity and tissue specificity. Here, we report a new strategy to construct SPs for efficient and muscle-specific gene expression. First, 19 nucleic acid motifs classified to 3 kinds of transcriptional regulatory elements were rationally selected. A recombinant promoter library was constructed by randomly assembling these motifs. Second, the transcriptional activities of ~1200 SPs were screened by intramuscular expression of several reporter genes in different cell lines for activity higher than that of the cytomegalovirus (CMV) promoter, with SP-301 finally identified as the strongest. A single intramuscular injection of mice with an SP-301 plasmid expressing mouse growth hormone releasing hormone accelerated mouse growth significantly over 24 days. Third, the muscle specificity of SP-301 was confirmed in transgenic mice. Finally, in comparison with the CMV promoter, SP-301 accelerated translocation and increased the level of plasmid in the nuclei of myoblast cells to a greater extent than in non-muscle cells. Altogether, the study has provided a more rational strategy to construct efficient and tissue-specific promoters, with the promoter SP-301 exhibiting promising potential for establishing an intramuscular gene expression system for therapeutic applications.