Northern Blot Hybridizations of RNA from AD 5 Transformed Cell Lines

Abstract

Researchers have tried to identify the viral factor involved in making a transformed cell oncogenic in newborn hamsters. Although studies on the viral proteins from transformed cells failed to show a relationship to oncogenicity, this study intended to identify and map viral mRNAs from these transformed cell lines in order to test the same relationship. Northern blot hybridizations were used to study the AD5 E1 region specific mRNAs from ten transformed cell lines. These cell lines had been transformed with either virus, total restriction endonuclease digested viral DNA, or specific fragments of viral DNA from the left hand end of the genome. All of these lines were selected for their varying oncogenicity in newborn hamsters, and for the size of the transforming fragment. Ad5 DNA fragments inserted into recombinant plasmids were used as the probes for detection of mRNAs, providing tools for mapping transformed cell mRNAs to specific regions of the Ad5 genome. The results failed to show a relationship between E1 region mRNA production and oncogenicity, but did reveal unusual mRNA transcription patterns from most of the cell lines. All the transformed cell lines tested appeared to have only an E1B 22s mRNA as the major mRNA from that region. An E1A/E1B cotranscript was identified as well as viral/cell chimeric mRNAs. These chimeras were due to either run-off transcription or to preinitiation in cellular sequences.