A BACULOVIRUS (BOMBYX-MORI NUCLEAR POLYHEDROSIS-VIRUS) REPEAT ELEMENTFUNCTIONS AS A POWERFUL CONSTITUTIVE ENHANCER IN TRANSFECTED INSECT CELLS

It has been previously reported that baculovirus homologous regions, t he regions of baculovirus genomes that contain the origins of DNA repl ication, can augment the expression of a small number of baculovirus g enes in vitro, We are now reporting that a region of the genome of Bom byx mori nuclear polyhedrosis virus (BmNPV) containing the homologous region 3 (HR3) acts as an enhancer for the promoter of a nonviral gene , the cytoplasmic actin gene of the silkmoth B. mori. Incorporation of the HR3 sequences of BmNPV into an actin promoter based expression ca ssette results in an augmentation of transgene expression in transfect ed cells by two orders of magnitude relative to the control re combina nt expression cassette, This increase is due to a corresponding increa se in the rate of transcription from the actin promoter and not to rep lication of the expression cassette and occurs only when the HR3 eleme nt is linked to the expression cassette in cis, A comparable degree of enhancement in the activity of the silkworm actin promoter occurs als o in heterologous lepidopteran cells, Concomitant supplementation of t ransfected cells with the BmIE1 trans-activator, which was previously shown to be capable of functioning in vitro as a transcriptional co-ac tivator of the cytoplasmic actin gene promoter, results in more than a 1,000-fold increase in the level of expression of recombinant protein s placed under the control of the actin gene promoter, These findings provide the foundation for the development of a nonlytic insect cell e xpression system for continuous high-level expression of recombinant p roteins. Such a system should provide levels of expression of recombin ant proteins comparable to those obtained from baculovirus expression systems and should also have the additional advantage of continuous pr oduction in a cellular environment that, in contrast to that generated by a baculovirus infection, supports continuously proper posttranslat ional modifications of recombinant proteins and the capability of expr ession of proteins from genomic as well as cDNA sequences.