Ecdysone-induced activation of membrane protein genes during morphogenesis of [formula omitted] imaginal discs : [formula omitted]. [formula omitted], [formula omitted]. [formula omitted], and [formula omitted]. [formula omitted]. Department of Genetics, University of California, Berkeley, CA, 94720, U.S.A.

Abstract

Previously, we established a method to produce recombinant proteins (r-proteins) in cocoons of germline transgenic silkworms, and showed that a step(s) in post-transcription processes was rate-limiting in obtaining a high yield of r-proteins. In this study, we examined whether the 5′-untranslated region (5′-UTR) of the polyhedrin gene (pol) of nucleopolyhedrovirus (NPV) has a translational enhancer activity in the r-protein expression by middle silk gland (MSG) cells of silkworm Bombyx mori (Bm). Sericin 1 gene (ser1) promoter-driven transformation vectors were constructed in which pol5′-UTRs of NPVs isolated from four different species, Bm, Spodoptera frugiperda, Ectropis oblique, and Malacosoma neustria, were each placed upstream of a reporter gene. Transient expression assays in MSGs showed that these pol5′-UTRs all enhanced the protein expression of reporter genes, and the pol5′-UTR of Bm NPV (pol5′-UTR/Bm) was the most effective among them. Thus, transgenic silkworms were generated, which bore the ser1 promoter-driven His-tagged secretory EGFP (sEGFP-His) gene under the control of pol5′-UTR/Bm. The synthesis of sEGFP-His proteins in MSGs of the transgenic worms was approximately 1.5-fold higher than that in those bearing null vectors. However, its mRNA expression levels were 67% of the control worms, indicating that the pol5′-UTR/Bm specifically enhanced the translational level. In conclusion, pol5′-UTR/Bm increased the yield of r-protein production in transgenic silkworms by enhancing the translational activity and this 5′-UTR could be useful for the mass production of r-proteins in germline transgenic silkworms.