Simian virus 40 (SV40) strains with novel properties generated by replacing the viral enhancer with synthetic oligonucleotides

Typical enhancers of viral or cellular genes are approx. 100-400 bp long and contain several transcription factor binding sites. Previously we have shown that SV40 genomic DNA that lacks its own enhancer can be used as an "enhancer trap" since it re-acquires infectivity upon incorporation of heterologous enhancers. Here we show that SV40 infectivity can be restored with synthetic enhancers that are assembled by the host cell. We found that several oligonucleotides, co-transfected with enhancerless SV40 DNA into host cells, were incorporated into the viral genome via cellular DNA-end joining. The oligonucleotides tested included MREs (metal response elements), the binding site for the transcription factor MTF-1, which induces gene activity in response to heavy metals. These recombinant SV40 strains showed preferential growth on cells overloaded with zinc or cadmium. We also co-transfected enhancerless SV40 DNA with oligonucleotides corresponding to enhancer motifs of human and mouse cytomegalovirus (HCMV and MCMV). In contrast to SV40 wild type, the viruses with cytomegalovirus-derived patchwork enhancers strongly expressed T-antigen in human HEK293 cells, accompanied by viral DNA replication. Occasionally, we also observed the assembly of functional viral genomes by incorporation of bovine DNA fragments, an ingredient of the fetal calf serum in the medium. These fragments contained, among others, binding sites for AP-1 and CREB transcription factors. Taken together, our studies show that viruses with novel properties can be generated by intracellular incorporation of synthetic enhancer DNA motifs