FIGURE 3.

Base-pairing with DsrA induces RNase III cleavage in the rpoS 5′-UTR at an alternative site. (A) Primer extension analysis of total RNA isolated from the wild-type E. coli strain (wt) and isogenic RNase III (rnc), Hfq (hfq), DsrA (dsrA⁻), and double (rnc/dsrA⁻) mutants grown at 28°C and 37°C. The product of RNase III cleavage within the rpoS leader (position G₋₁₁₂) accumulates only in the wild-type strain. (B) In vitro synthesized DsrA was incubated alone or with RNase III in the absence (−) or presence (+) of equimolar amount (1:1), fourfold, or ninefold molar excess of RpoSI for 10 min and further analyzed by primer extension. The arrow indicates the nucleotide (A₂₈) at which RNase III cleaves DsrA complexed with RpoSI. (C) Depicted is a part of the rpoS/DsrA complex. Arrows indicate the internucleotide bonds that are cleaved by RNase III. The coordinates of nucleotides in close vicinity to the scissile bonds (G₋₁₁₂ and A₂₉) were determined by primer extension (see Figs. 2B, 3B, respectively).