Regulation of circadian clock gene expression by phosphorylation states of KaiC in cyanobacteria.
Three clock proteins--KaiA, KaiB, and KaiC--have been identified as essential components of the circadian oscillator in cyanobacteria, and Kai-based chemical oscillation is thought to be the basic circadian timing mechanism in Synechococcus elongatus PCC 7942. Transcription and translation of kaiBC in cyanobacterial cells was quantitatively studied to elucidate how these processes are coupled to the chemical oscillator using a strain in which circadian oscillation is under the control of IPTG (isopropyl-beta-D-thiogalactopyranoside). The kinetics of repression of kaiBC promoter triggered by IPTG allowed estimation of transient response at 10 h. This response time is suitable for cyanobacterial transcription and/or translation to match with the Kai-based oscillator. Interestingly, kaiBC promoter activity and KaiC phosphorylation showed robust circadian rhythms, whereas trc promoter-driven kaiBC mRNA levels and KaiC accumulation were almost arrhythmic. These results indicate that cyanobacterial circadian rhythms can be generated even if kaiBC expression is constitutive. Moreover, there was a positive correlation between activation of the kaiBC promoter and an increase in the KaiC phosphorylation ratio in three rhythmic conditions. Based on these observations, it is likely that the KaiC phosphorylation ratio is the main factor in the activation of kaiBC promoter. Finally, we quantitatively compared the threshold level of phosphorylated KaiC for the repression or derepression of kaiBC promoter and found that this parameter is an important factor in repressing the kaiBC promoter.
Murayama
Yoriko
Y
Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho 1, Chikusa-ku, Nagoya 464-8602, Japan.
Oyama
Tokitaka
T
Kondo
Takao
T
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D002940:Circadian Rhythm / Q000235:genetics*; D056925:Circadian Rhythm Signaling Peptides and Proteins; D015964:Gene Expression Regulation, Bacterial; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D007700:Kinetics; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D046940:Synechococcus / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D056925:Circadian Rhythm Signaling Peptides and Proteins; C114269:KaiA protein, cyanobacteria; C114270:KaiB protein, cyanobacteria; C114271:KaiC protein, cyanobacteria; D012333:RNA, Messenger; D007544:Isopropyl Thiogalactoside