Overexpression of the adenovirus type 12 (Ad12) pTP or E1A gene facilitates Ad12 DNA replication in nonpermissive BHK21 hamster cells

In the adenovirus type 12 (Ad12) hamster cell system, abortive virus infect ion is one of the factors associated with the highly efficient oncogenesis in newborn Syrian hamsters. We have shown earlier that the replication and efficient late transcription of the Ad12 genome are blocked in Syrian hamst er cells. Some of the early Ad12 functions are transcribed in these cells, although at a minimal rate. In the present study, we demonstrate that low e xpression levels of the E1A and precursor to terminal protein (pTP) genes o f Ad12 seem to be responsible for the lack of Ad12 DNA replication in hamst er cells. The Ad12 genes for the E1A functions or for pTP were tethered to the strong early promoter of the human cytomegalovirus and transfected into BHK21 cells. Subsequently, these cells were infected with Ad12 virions. In Ad12-infected BHK21 cells, which overexpressed pTP or E1A, full-length Ad1 2 DNA was de novo synthesized, as documented by metabolic labeling with [H- 3]thymidine and by zone velocity sedimentation in alkaline sucrose gradient s followed by gel electrophoresis of the H-3-labeled DNA and Southern blot hybridization to (32)p-labeled Ad12 DNA. Transfection of the cloned E1A reg ion of Ad2 yielded similar results. The newly synthesized Ad12 DNA was cova lently linked to pTP. The Ad12 DNA binding protein (DBP) and DNA polymerase (pol) genes were transcribed at levels similar to those in merely Ad12-inf ected cells. In pTP or E1A gene-transfected and Ad12-infected BHK21 cells, marginal levels of late Ad12 mRNA were detectable. Late Ad12 proteins were, however, not synthesized. Apparently, Ad12 DNA replication in hamster cell s is rendered impossible due to insufficient threshold levels of the viral E1A and/or pTP.