2 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 substra tes ATP and CTP for stabilizing the enzyme-DNA association (Gourse, R. L. (1988) Nucleic Acids Res. 16, 9789-9809). We show that in the abse nce of UTP and GTP the stabilization is accomplished by short RNA olig omers synthesized in an unusual ''-3-->'' mode whereby the primer init iated at the +1 site presumably slips back by three nucleotides into t he -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 readil y aborted from the promoter complex. The stable -3--> ternary complexe s carry sigma factor but otherwise resemble elongation complexes in th eir high salt stability and in the fact that they are formed with a mu tant RNA polymerase deficient in promoter binding. A model is proposed explaining the stability of the -3--> ternary complexes by RNA slippi ng into a putative ''tight RNA binding site'' in RNA polymerase which is normally occupied by RNA during elongation.