Two modes of transcription initiation in vitro at the rrnB P1 promoter of Escherichia coli

The rrnB P1 promoter of Escherichia coli (starting sequence C^-4-A^-3-C^-2-C^-1-A ⁺¹-C⁺²-U⁺³-G⁺⁴) forms a binary complex with RNA polymerase that is highly unstable and requires the presence of transcription substrates ATP and CTP for stabilizing the enzymeDNA association (Gourse, R. L. (1988) Nucleic Acids Res. 16, 9789-9809). We show that in the absence of UTP and GTP the stabilization is accomplished by short RNA oligomers synthesized in an unusual "-3→ " mode whereby the primer initiated at the +1 site presumably slips back by three nucleotides into the -3 site and is then extended yielding stable ternary complexes. By contrast, short oligomers initiated in the conventional "+1→" mode without slippage do not exert the stabilization effect and are readily aborted from the promoter complex. The stable -3→ ternary complexes carry σ factor but otherwise resemble elongation complexes in their high salt stability and in the fact that they are formed with a mutant RNA polymerase deficient in promoter binding. A model is proposed explaining the stability of the -3→ ternary complexes by RNA slipping into a putative "tight RNA binding site" in RNA polymerase which is normally occupied by RNA during elongation.