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Promoter determinants for Escherichia coli RNA polymerase holoenzyme containing σ 38 (the rpoS gene product)

1995; Oxford University Press; Volume: 23; Issue: 5 Linguagem: Inglês

10.1093/nar/23.5.827

1362-4962

Kan Tanaka, Shuichi Kusano, Nobuyuki Fujita, Akira Ishihama, Hideo Takahashi,

Bacteriophages and microbial interactions

Sequence determinants responsible for promoter recognition by RNA polymerase holoenzyme containing σ 38 , the rpoS gene product, were analyzed. In a previous study [Tanaka et al . (1993) Proc. Natl. Acad. Sci. USA , 90, 3511-3515], Escherichia coli promoters were classified into three groups: promoters recognized only by RNA polymerase holoenzyme containing σ 70 (Eσ 70 ); promoters recognized preferentially by that containing σ 38 (Eσ 38 ); promoters recognized by both Eσ 70 and Eσ 38 . As representatives of each group of promoter, we chose the alaS, fic and lac UV5 promoters. Making use of a restriction enzyme site inserted between the −10 and −35 hexamer sequences, promoters were divided into the upstream (UE) and downstream (DE) elements. These UEs and DEs were combined in all possible combinations and used for in vitro transcription reactions. Promoters containing DE from the fic or lac UV5 promoter were found to be recognized by Eσ 38 , while those containing DE from the alaS promoter were not. Moreover, fic DE alone functioned as an efficient promoter for Eσ 38 . Thus we conclude that the discrimination signal resides within the DE sequence. To test the activator response of Eσ 38 , in vitro transcription reactions were also performed with the gal and lac promoters. For both CRP-responsive P1 promoters, Eσ 38 was found to be activated by the CRP-cAMP complex.