Totally asymmetric transcription of coliphage T7 in vivo: Correlation with poly G binding sites

Abstract

In vivo transcription of T7 phage was studied by means of hybridization between phage-specific radioactive RNA and separated complementary strands of T7 DNA. The latter were preparatively separated by CsCl density-gradient centrifugation in the presence of guanine(G)-rich polyribonucleotides, which bind to one strand only. The separated strands failed to renature when annealed separately but renatured when mixed together. Pulse-labeled 3H-RNA, isolated from cultures of Escherichia coli B at various times after T7 infection, hybridized exclusively with the poly G-binding strand H, indicating that only this strand is transcribed in vivo. Non-poly G-binding strand L did not hybridize (less than 0.02%) with the in vivo synthesized T7 RNA, but both the H and L strands hybridized with enzymatically prepared 3HRNA transcribed from denatured T7 DNA template. The fact that the poly G-binding, deoxycytidylate (dC)-rich clusters are restricted to the in vivo transcribing H strand, together with the absence of thymine-rich clusters in T7 DNA, is compatible with the hypothesis (Szybalski et al., 1966) that pyrimidine-rich clusters are related to RNA transcription, possibly as the initiation and termination sites.