Animal DNA‐Dependent RNA Polymerases

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Abstract
The RNA synthesized in vitro on superhelical Simian virus 40 (SV40) DNA (form I) by calf thymus and rat liver RNA polymerases B and by calf thymus RNA polymerase AI was characterized. Both 32P‐labeled ATP and GTP were found at the 5′ end of the RNAs synthesized by AI and B enzymes indicating that each type of enzyme possesses at least two initiation sites on SV40 DNA form I. Competition experiments between AI and B enzymes for initiation sites on SV40 DNA form I suggest that some of these sites could be different for the two enzymes. The chain growth rates of RNA chains synthesized by calf thymus AI and B enzymes on SV40 DNA form I were determined. Both calf thymus RNA polymerase AI and calf thymus and rat liver RNA polymerases B transcribed symmetrically SV40 DNA form I under a variety of incubation conditions. Under similar conditions Escherichia coli RNA polymerase holoenzyme transcribed asymmetrically SV40 DNA form I, while symmetrical transcription was observed with E. coli core enzyme. The maximum size of the RNAs synthesized by calf thymus enzyme AI was several times the length of the SV40 genome, indicating that this enzyme can pass several times over its initiation site and transcribe SV40 DNA form I over the entire genome. In contrast, RNAs made by enzymes B had a more definite maximum size (18–20 S), indicating differences between AI and B enzymes at the level of chain termination. However, this shorter size of RNA's synthesized by enzymes B does not correspond to a selective transcription of some part of the viral genome, since our results indicate that most, if not all, of the SV40 genome was transcribed by enzymes B. The arrest of elongation of RNA chains synthesized by enzymes B was related to the low ionic strength incubation conditions and did not reflect a true termination step. It appears therefore that, when transcribing SV40 DNA form I in vitro, mammalian RNA polymerases AI and B do not exhibit the two main characteristics of the prokaryotic transcription, namely asymetry and selectivity. Our results are discussed in the light of the present knowledge of the transcription of SV40 DNA in vivo during the lytic cycle.