| The E.coli DnaA protein is the replication initiator protein, it is also a transcription factor. The LexA protein regulates the SOS response. The function of ybfE gene remains elusive. Expression of ybfE is suggested to be regulated by LexA. DnaA and LexA regulate gene expression by their bindings to their conservative sequences in promoters and these binding sites are named DnaA-box and LexA-box. There are six DnaA-box and one LexA-box in the ybfE promoter.The paper investigated roles of the DnaA and LexA protein in regulation of the ybfE gene expression. The lacZ gene was inserted into downstream of the ybfE promoter on chromosome. In order to analyze the effect of DnaA or LexA on ybfE expression, expression levels of the lacZ reporter gene in different availability of DnaA or LexA were tested by Miller method. The results showed that there were no significant changes in ybfE expression when DnaA was overexpressed or reduced by titration of DnaA to the datA site. In dnaA345 mutant cells, the activity of ybfE promoter was obviously higher than that of wild type cells. In lexA3 mutant cells and â–³lexA cells, the activity of ybfE promoter had no obvious change compared with the wild-type cells. And then, we inserted the native ybfE promoter in front of the promoterless lacZ gene into pTAC3953 plasmid. The expression of lacZ gene was then measured in different availability or activity of DnaA, LexA. Also, after the deletion or the mutation of DnaA-box or LexA-box in pTACYL plasmid, the expression of lacZwas measured. The results showed that the expression of ybfE was not significantly changed by mutations in DnaA-box, but mutations in LexA-box significantly declined expression of ybfE. Mutations in LexA-box increased expression of the ybfE gene in dnctA345 cells. A gel-shift experiment showed that the LexA protein bound to the ybfE promoter. These results suggest that the ybfE gene expression is likely to be regulated by the DnaA and LexA protein. |
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