Strand Orientation of Simian Virus 40 Transcription in Productively Infected Cells

Abstract

Strand orientation of transcription in BSC-1 cells infected by simian virus 40 (SV40) was investigated by annealing RNAs extracted from infected cells with asymmetric complementary RNA from SV4O DNA synthesized in vitro by RNA polymerase from Escherichia coli. The results suggest that the early viral RNA sequences (those made before replication of viral DNA) and the late RNA sequences (those synthesized concurrently with replication of viral DNA) are transcribed from opposite strands of the SV40 DNA. The RNA synthesized in vitro is of the same polarity as the early in vivo RNA and is complementary to the late in vivo RNA. Knowledge of strand selection in lytic infection is a step towards understanding the regulation of transcription of this viral DNA.The small DNA virus, simian virus 40 (SV40), grows lytically in permissive African green monkey cells and usually transforms nonpermissive cells, such as mouse or hamster, in which a complete replicative cycle is absent. This difference of interaction with host cells often correlates with a difference in transcription of the viral genome. In the lytic cycle about one-third to one-half of the genome is transcribed early in infection, before replication of viral DNA begins, and comprises the class of early RNA(s) (1-5). After DNA replication has begun, the remainder of the genome (1, 6) is transcribed such that late in infection both early and late RNA sequences are synthesized. In certain transformed cells, transcription is limited to about 40% of the genome (1, 2). Variable amounts of the genome are transcribed in other transformed lines (6).Most of the sequences transcribed in transformed cells are also transcribed early in lytic infection (2, 4).To understand how the various modes of transcription might be regulated, we have investigated which strand of viral DNA serves as template for RNA synthesis in vivo. This was made possible by the observation that, in vitro, Escherichia coli RNA polymerase preferentially transcribes only one DNA strand, often in its entirety (7). We compared viral RNA made early and late in infection with that made in vitro. The results show that early sequences are of the same polarity as the in vitro RNA. whereas the late sequences are complementary to the in vitro RNA. We conclude that early and late viral RNA species are transcribed from different strands of the SV40 DNA. (11)