| Research Abstract |
We transferred a genomic DNA library into circadian clock mutants of Synechococcus sp. Strain PCC 7942, selected transformed clones with kanamycin, screened cured clones which showed wildtype phenotype, and recovered the transferred genomic DNA segments from cured clones into E.ccli cells as plasmids by the plasmid rescue procedure. We identified a gene cluster kaiABC which is composed of three genes, kaiA, kaiB, and kaiC and is essential to the clock function (Ishiura et al., 1998. Science 281 : 1519-1523). All the mutations analyzed were missense mutations and mapped to this gene cluster. Homologs of kaiB and kaiC were found in the genomes of archaea, but not found in eukaryotes. Two promoters(PkaiA and PkaiBC), each one in both the upstream regions of kaiA and kaiB, were found, and kaiA and a set of kaiB and kaiC were transcribed as a kaiA mRNA and a kaiBC mRNA, respectively. The expression of both kaiA and kaiBC was circadian and affected by mutations in kaiA, kaiB, and kaiC themselves. KaiA protein enhanced the activity of PkaiBC promoter whereas KaiC protein repressed the activity. Therefore, the essence of the circadian clock in cyanobacteria is the circadian autoregulation of the expression of kaiABC by Kai proteins, KaiA, KaiB, and KaiC.
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