Role of CspC and CspE in Regulation of Expression of RpoS and UspA, the Stress Response Proteins in Escherichia coli

When exponentially growing Escherichia coli cells are transferred to a low temperature of 15°C, there is a growth lag period, during which the synthesis of cold shock proteins is induced transiently. Among these, CspA is a major cold shock protein. The CspA family of E. coli comprises nine homologous proteins, CspA to CspI. These are of similar sizes, and the corresponding genes are scattered on the E. coli chromosome (for reviews, see references 26, 28, and 37). Of these proteins, only CspA, CspB, CspG, and CspI are cold shock inducible. However, their induction patterns are different. Maximum induction of CspA is at 10 to 24°C, while that of CspB and CspG is at 15°C (7). CspI is produced at 10 to 15°C (36). CspD is induced upon nutritional starvation (38). CspC and CspE are produced mainly at 37°C. These two proteins were originally identified as multicopy supressors of a temperature-sensitive chromosome-partitioning mutant (39). CspE is constitutively produced throughout the growth stages except the lag period, when a 1.5-fold increase in its production is observed (3). It has been shown that CspE inhibits phage lambda Q-mediated transcriptional antitermination in vitro (11). CspE has also been shown to negatively regulate the expression of CspA by increasing the promoter-proximal pausing efficiency of the RNA polymerase (3). CspA has been proposed to act as an RNA chaperone that facilitates translation at low temperature by blocking the formation of secondary structures in mRNA (16). E. coli CspA-family RNA chaperones are shown to be transcription antiterminators (4). CspA has 43% identity to the cold shock domain of the eukaryotic Y-box protein family. This protein family is implicated in various cellular functions such as transcription, DNA replication and repair, and masking of maternal mRNAs (5, 30). CspA binds to RNA with low sequence specificity and low binding affinity. In contrast, CspB, CspC, and CspE are able to selectively bind RNA and single-stranded DNA sequences (27). However, it remains to be seen if this selectivity has any in vivo significance. In spite of the recent extensive studies (for reviews, see references 26, 28, and 37) on the CspA family of E. coli, the cellular functions of these proteins are not fully elucidated and it is not clear why E. coli has so many CspA-like proteins. Yamanaka et al. (37) had suggested that this large Csp family probably originated from a number of gene duplications and, after subsequent adaptation, resulted in specific groups of genes responding to different environmental stresses. In the present study, using two-dimensional gel electrophoresis, the effect of overproduction of CspC and CspE on the overall protein pattern of the cell was tested. The proteins that were significantly and consistently upregulated by the overexpression of CspC and CspE were identified. This result was analyzed by studying the effect of CspC on the amount and stability of mRNAs for these proteins by primer extension. Interestingly, these three proteins, OsmY, Dps, and UspA, are induced in response to different stresses and also upon stationary phase (2, 13, 14, 17, 24). RpoS is a regulatory element for osmY and dps (19, 42). Further analysis showed that the amount and stability of the rpoS mRNA, as well as the level of RpoS itself, are greatly enhanced by the overexpression of CspC and CspE and that this may result in the upregulation of OsmY and Dps. This possibility was confirmed by using the rpoS deletion strain. The other RpoS-regulated proteins such as ProP and KatG were also upregulated by the overexpression of CspC. The significance of this observation for a possible role of CspC and CspE in the regulation of expression of stress response proteins is discussed.