TRANSCRIPTIONAL REGULATION OF CELL LINE-DEPENDENT, BACULOVIRUS-MEDIATED EXPRESSION OF FOREIGN GENES

Cell line-dependent expression of foreign genes in the baculovirus sys tem was investigated using a recombinant vAc beta hCG-luc virus carryi ng two reporter genes-beta subunit of human chorionic gonadotropin (be ta hCG) and luciferase (luc)-placed under the transcriptional control of the Autographa californica nuclear polyhedrosis virus (AcNPV) polyh edrin gene promoters, Five different lepidopteran cell lines derived f rom Spodoptera frugiperda (Sf21 and Sf9). Bombyx mori (BmN and Bm5), a nd Trichoplusia ni (TN368) were used as host cells, TN368 expressed bo th beta hCG and LUC to maximum levels, followed by BmN, Sf21, and Sf9 in descending order, Bm5 did not show any evidence of synthesis of the two proteins. Dot blot analysis of DNA from the vAc beta hCG-luc-infe cted cells revealed that the level of entry of viral DNA was the same for all the five cell lines, After the completion of viral DNA replica tion (18 hr post infection), the level of viral DNA was the same for a ll the cell lines except for Bm5 where viral DNA replication did not t ake place and the residual virus was cleared from the cells. Analysis of RNA from the four expressing cell lines revealed a direct correlati on between protein levels and levels of mRNA, suggesting transcription al control. Differences in mRNA stability between cell lines was also evident. Gel retardation analysis of a host factor binding to transcri ptionally important sequence motifs within the AcNPV polyhedrin gene p romoter revealed an inverse correlation between the levels of this pol yhedrin promoter-binding protein (PPBP) and reporter gene expression, Cold competition and mutation analyses of the DNA-protein complexes in dicated that PPBP present in different cell lines recognized the same DNA sequence motifs present within the polyhedrin promoter.