Osmo-Regulation of Bacterial Transcription via Poised RNA Polymerase Shun Jin Lee, Jay D Gralla  Molecular Cell  Volume 14, Issue 2, Pages 153-162 (April 2004) DOI: 10.1016/S1097-2765(04)00202-3

Figure 1 Single-Round In Vitro Transcription Experiments Measuring the Effect of Increasing Ionic Solute Concentrations (A) Potassium glutamate titration. osmY and fic-con promoters were transcribed by σ38 and the lacUV5 promoter by σ70 in a heparin-challenge protocol. Potassium glutamate concentrations were 100 mM (lanes 1, 5, and 9), 200 mM (lanes 2, 6, and 10), 300 mM (lanes 3, 7, and 11), and 400 mM (lanes 4, 8, and 12), and transcription was for 5 min. Percent transcription was calculated by standardizing to the highest signal for each set of experiments. White bars represent 100 mM, light gray 200 mM, gray 300 mM, and black 400 mM potassium glutamate. (B) Percent transcription versus time at 100 mM and 400 mM potassium glutamate at the osmY promoter. (C) Potassium chloride titration. Potassium chloride concentrations were 100, 200, 300, and 400 mM from left to right for each experiment. Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)

Figure 2 Footprinting of σ38 RNA Polymerase on Supercoiled Plasmid DNA (A) KMnO4 footprinting at the osmY promoter. (B) DNase I footprinting of osmY and fic-con. Gray brackets on the left indicate the region of hypersensitive sites detected at osmY. Black brackets indicate the protected region for both osmY and fic-con. Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)

Figure 3 Multiple-Round Transcription of the osmY Promoter Left panel, 100 mM potassium glutamate. Right panel, 400 mM potassium glutamate. The time points were 2.5 min (lanes 1 and 5), 5 min (lanes 2 and 6), 10 min (lanes 3 and 7), and 15 min (lanes 4 and 8). The time to complete a single round of transcription was determined by single-round time course experiments and was ∼3 min for 100 mM potassium glutamate and ∼5 min for 400 mM potassium glutamate. Relative transcription versus time is plotted. Relative transcription is the amount of transcription normalized to single-round transcription levels at 100 mM potassium glutamate (e.g., relative transcription of 6 indicates that six times as much transcript is produced as compared to the amount produced during a single round of transcription at 100 mM potassium glutamate). Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)

Figure 4 Single-Round Transcription Experiment of Various osmY G-Element Mutants at Increasing Potassium Glutamate Concentrations Potassium glutamate concentrations were 100 mM (lanes 1, 5, and 9), 200 mM (lanes 2, 6, and 10), 300 mM (lanes 3, 7, and 11), and 400 mM (lanes 4, 8, and 12). Percent transcription was calculated by standardizing the signal to osmY transcription levels at 300 mM potassium glutamate. White bars represent 100 mM, light gray 200 mM, gray 300 mM and black 400 mM potassium glutamate. Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)

Figure 5 Single-Round Transcription Experiments Measuring the Effect of Various Electro-Neutral Osmolytes on σ38- and σ70-Dependent Transcription osmY and fic-con promoters were transcribed by σ38 and the lacUV5 promoter by σ70. The levels of trehalose used (top panels) were, from left to right, no trehalose, 100 mM, 200 mM, 300 mM, and 400 mM. The levels of L-proline (middle panels) were, from left to right, no L-proline, 100 mM, 200 mM, 300 mM, and 400 mM. The levels of glycine betaine (bottom panels) were no glycine betaine, 100 mM, 200 mM, 300 mM, 400 mM, 500 mM, and 700 mM. Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)

Figure 6 Model for How Escherichia coli Changes Its Transcription in Response to an Osmotic Shock Molecular Cell 2004 14, 153-162DOI: (10.1016/S1097-2765(04)00202-3)