Punctuation of transcription in vitro of the tryptophan operon of Escherichia coli

Abstract

RNA transcribed in vitro from DNA of a tryptophan (trp) transducing strain of bacteriophage ϕ80 which contains the trp regulatory elements consists of a polycistronic messenger transcribed from the structural genes, and possibly the regulatory region, and a separate RNA species (called trp regRNA) which is transcribed from the regulatory region. This conclusion is based on hybridization experiments with trp RNA synthesized in vitro and the separate DNA strands of trp transducing strains of λ with and without the trp regulatory elements. The length of trp regRNA determined by filtration on Sephadex G-200 is 110–180 nucleotides. From the amount and the length of trp regRNA we have calculated that 8–20 copies of trp regRNA are synthesized per copy of polycistronic trp mRNA. We conclude that during transcription of the trp operon RNA polymerase frequently is rejected at a specific site ahead of the first structural gene, trpE. The termination factor Rho is not involved in this process. A different protein fraction, which specifically stimulates the synthesis of trp enzymes in an in vitro protein-synthesizing system (Pouwels and Van Rotterdam, 1975), was found to antagonize the abortive synthesis of trp mRNA. A model is proposed for the control of transcription of the trp genes, which operates through a mechanism of punctuation of RNA synthesis at a specific site on the DNA template and anti-termination of RNA synthesis by means of a positive control factor.