| Abstrakt: |
Transcription of the psbA2 gene in the unicellular photosynthetic cyanobacterium Microcystis aeruginosa K-81 is modulated by light and follows a circadian rhythm. In this study, we further characterized psbA transcription using a series of 5′-upstream deletions and mutant promoters which were tested in both photosynthetic and non-photosynthetic bacteria. Specific psbA2 transcripts were obtained from a minimal promoter sequence (–38/+14) with Escherichia coli RNA polymerases (RNAPs) both in vivo and in vitro, indicating the presence of a common regulatory mechanism for basal transcription. A DNase I footprinting assay showed that the E. coli RNAP, which is structurally similar to that of cyanobacteria, specifically binds to a large segment (from –115 to +23) of the sequence upstream of psbA2. In cyanobacteria, the –10 sequence (TAGTAT), but not the –35 motif (TTTACA), is essential for basal transcription by homologous and heterologous RNAPs that contain the major sigma factor. Each of the conserved thymidine nucleotides at positions –12 and –7 (underlined above) was essential, and both an insertion and a deletion in the spacer region of the promoter caused reductions in transcription. RNAP was able to bind to a mutant promoter lacking the –10 sequence, though this did not actually lead to transcription. Interestingly, a high level of arrhythmic circadian transcription was observed in mutants lacking the –35 region. In contrast, a mutation in the AU-box mutation, which controls the stability of the psbA2 mRNA, did not affect the circadian pattern of transcription. These findings demonstrate that light-dependent psbA2 expression is controlled at the transcriptional and post-transcriptional levels, whereas the circadian pattern of expression is regulated at the transcriptional level. [ABSTRACT FROM AUTHOR] |
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