Identification of Promoters in the Rhizobium etli Genome.


All bacteria, except Deinococcus radiodurans, contain a main sigma subunit, also known as housekeeping or sigma 70 ( s 70 ), which controls the expression of most of the genes in any growth condition. The s 70 subunit of E. coli and Bacillus subtillis recognize two types of promoter sequence consensus: the first one, or “canonical promoter”, consists of two hexamer sequences [TTGACA] and [TATAAT], centered respectively in the positions -10 and -35 relative to the transcription start site. The second one, named “extended–10 promoter” consist of an element [TATAAT] plus two nucleotides [TG], located in the –12 and –13 positions. In this promoter type a –35 region or equivalent sequences does not exist. Thought these promoter systems have been thoroughly studied in E. coli and B. subtillis, nothing is known about the promoter structure in the majority of the bacteria. Rhizobium etli possesses a housekeeping s 70 (SigA) larger than the E. coli s 70 . Most of the s 70 promoters already characterized in R. etli are not transcribed by the E. coli RNApol in vivo or in vitro. In contrast, the RNApol holoenzyme of R. etli can initiate transcription of typical E. coli s 70 promoter. This observation suggests some differences between the transcriptional machinery of E. coli and R. etli , perhaps at the level of promoter recognition by s 70 factor. To analyze the molecular basis of the R. etli gene expression, in this study we want to identify, characterize and sequence active promoters of R. etli under exponential growth conditions.

Participants:

Dr. Miguel Ramírez, experimental design, performing experiments and data analysis.
C. Gamaliel López Leal, promoter analysis by saturation mutagenesis.
Dr. Guillermo Dávila, data analysis.


Evolutionary Genomics Program