Role of CspC and CspE in regulation of expression of RpoS and UspA, the stress response proteins in Escherichia coli - PubMed (original) (raw)

FIG. 3

Stability and amount of the osmY, dps, and uspA mRNAs in the wild-type, wild-type overexpressing CspC, and Δ_cspC_ Δ_cspE_ strains. The respective cultures were grown at 37°C to an OD600 of 0.5. Total RNA was extracted by the hot-phenol method. For measuring the stability of the mRNAs, rifampin was added at a final concentration of 200 μg ml−1 to stop the transcription. Samples (1.5 ml) were removed at each time point, RNA was isolated, and primer extension analysis was carried out with oligonucleotides corresponding to osmY, dps, or uspA. (A) For osmY mRNA, the wild-type (Wt) strain is lanes 1 to 4 (0, 10, 15, and 20, min, respectively), the Δ_cspC_ Δ_cspE_ strain is in lanes 5 to 7 (0, 8, and 10 min, respectively), and the wild-type strain overexpressing CspC is in lanes 8 to 10 (0, 20, and 30 min, respectively). For dps mRNA, the wild-type strain is in lanes 1 to 4 (0, 1, 2, and 4 min, respectively), the Δ_cspC_ Δ_cspE_ strain is in lanes 5 to 8 (0, 0.5, 1, and 2 min, respectively), and the wild-type strain overexpressing CspC is in lanes 9 to 12 (0, 4, 8, and 10 min, respectively). For uspA mRNA, the wild-type strain is in lanes 1 to 3 (0, 1, and 2 min, respectively), the Δ_cspC_ Δ_cspE_ strain is in lanes 4 to 7 (0, 0.5, 1, and 2 min, respectively), and the wild-type strain overexpressing CspC is lanes 8 to 11 (0, 10, 12, and 15 min, respectively). For ompF mRNA, the wild-type strain is in lanes 1 to 3 (0, 5, and 10 min, respectively), the Δ_cspC_ Δ_cspE_ strain is in lanes 4 to 6 (0, 5, and 10 min, respectively), and the wild-type strain overexpressing CspC is in lanes 7 to 9 (0, 5, and 10 min, respectively). (B) Graphical representation of the results in panel A, which shows the half-lives of the osmY, dps, uspA, and ompF mRNAs. The respective mRNA at the zero time point was takes as 100% in each case. Solid squares, mRNA (i.e., osmY, dps, uspA, or ompF) from cells overexpressing CspC; open squares, mRNAs from wild-type cells; open circles, mRNAs from Δ_cspC_ Δ_cspE_ cells.