Characterization of Escherichia coli cspE, whose product negatively regulates transcription of cspA, the gene for the major cold shock protein.
Escherichia coli contains nine members of the CspA protein family from CspA to Cspl. To elucidate the cellular function of CspE, we constructed a delta cspE strain. CspE is highly produced at 37 degrees C. The synthesis level of CspE transiently increased during the growth lag period after dilution of stationary-phase cells into the fresh medium at 37 degrees C. This is consistent with the delta cspE phenotype of the longer growth lag period after dilution. The protein synthesis patterns of the delta cspE strain and the wild-type strain were compared using two-dimensional gel electrophoresis. In the delta cspE strain, the synthesis of a number of proteins at 37 degrees C was found to be altered and cspA was derepressed. The derepression of cspA in the delta cspE strain was at the level of transcription in a promoter-independent fashion but was not caused by stabilization of the cspA mRNA, which was shown to be a major cause of CspA induction after cold shock. In vitro transcription assays demonstrated that both CspE and CspA enhanced transcription pause at the region immediately downstream of the cold box, a putative repressor binding site on the cspA mRNA. In a cell-free protein synthesis system using S-30 cell extracts, CspA production was specifically inhibited by the addition of CspE. These results indicate that CspE functions as a negative regulator for cspA expression at 37 degrees C, probably by interacting with the transcription elongation complex at the cspA cold box region.
Bae
W
W
Department of Biochemistry, Robert Wood Johnson Medical School, Piscataway, NJ 08854, USA.
Phadtare
S
S
Severinov
K
K
Inouye
M
M
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D015139:Blotting, Southern; D002455:Cell Division; D002474:Cell-Free System; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D003720:Densitometry; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D008957:Models, Genetic; D016296:Mutagenesis; D011401:Promoter Regions, Genetic; D013696:Temperature; D013997:Time Factors; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000378:metabolism
D001426:Bacterial Proteins; C088097:CspE protein, E coli; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins; C062260:cold shock protein CS7.4, Bacteria; C078564:universal stress protein A, Bacteria; D012321:DNA-Directed RNA Polymerases; D001616:beta-Galactosidase