Secretory pathway limits the enhanced expression of classical swine fever virus E2 glycoprotein in insect cells

The 3' untranslated region (UTR) is an important element that determines th e level of recombinant protein expression via baculovirus vectors. Previous work using chloramphenicol acetyl transferase as reporter has shown that p 10-promoter based baculovirus vectors with the authentic p10 3' UTR resulte d in higher expression levels than vectors carrying an SV40 early terminato r, as part of a lacZ selection cassette. To examine whether a similar incre ase in expression levels could be obtained for baculovirus-expressed glycop roteins, the classical swine fever virus E2 antigen was used as a model. Wi th the authentic p10 3' UTR higher levels of E2 transcript were found than in the presence of the SV40 terminator. This higher number of transcripts w as accompanied by elevated levels of intracellular, non-glycosylated E2 pro tein. However, the levels of intracellular glycosylated forms of E2 and of extracellular E2 were similar for both type of terminators. These results s how that translation of the recombinant mRNA is not the rate limiting step in the expression of glycoproteins, but the downstream processing and secre tion of the translation products. (C) 2001 Elsevier Science B.V. All rights reserved.