Promoter recognition by a cyanobacterial RNA polymerase: in vitro studies with the Calothrix sp. PCC 7601 transcriptional factors RcaA and RcaD.

To study the transcriptional apparatus and the mechanisms that control gene expression in cyanobacteria, the RNA polymerase was purified from the filamentous Calothrix sp. PCC 7601 and used in in vitro transcription assays. Conditions required for specific transcription initiation to occur were analyzed with the eleven Calothrix PCC 7601 genes for which the 5' ends have been mapped. Most of the transcripts directly obtained did not have the expected size, providing a test for looking at specific transcription factors. Addition of RcaA, a protein that binds to the promoter region of the phycobiliprotein cpeBA operon, restored accurate initiation of transcription in the in vitro system for three phycobiliprotein promoters. RcaA thus is a transcription factor that allows to mimick in vivo transcription. In parallel, the functional properties of the Escherichia coli and cyanobacterial RNA polymerases were compared. The enteric enzyme could not precisely initiate transcription at the promoter of a phycobiliprotein gene and, reciprocally, the cyanobacterial RNA polymerase could initiate transcription at PlacUV5, but not from wild-type Plac promoters. The different behaviours of the enzymes are discussed in the light of the structural differences that exist between subunits of the RNA polymerases.