Figure 1.

Thermosensing activity of CssA thermometer. (A) Proposed RNA secondary structure for the CssA RNA thermometers and for two variants (CssA1 and CssA2) prepared to perform nuclear magnetic resonance (NMR) studies; predicted structures were generated with VARNA (22). The lower panel shows the thermoregulated activity of the corresponding RNA constructs as assayed in Escherichia coli by western blot analysis of the protein products. The A44 and G45 nucleotides are highlighted with blue fill color on the CssA RNA secondary structure. (B) Selective 2′-OH acylation analyzed by primer extension (SHAPE) (21) for the CssA RNA establishes the RNA secondary structure at different temperatures, well below and just above activation of protein synthesis (4–42°C). (C) An overlay of the 1D imino region for the CssA RNA and for the two modified variants (CssA1 and CssA2). The considerable similarities in the imino resonances peak demonstrate the presence of very similar structures across all three RNAs. (D) Temperature-dependent SHAPE data summarized on the secondary structure of the CssA RNA; the 5′- and 3′-tails are reactive at all temperatures.