Regulation of circadian clock gene expression by phosphorylation states of KaiC in cyanobacteria
Regulation of circadian clock gene expression by phosphorylation states of KaiC in cyanobacteria
复制标题
DOI：
10.1128/jb.01693-07
复制
发表时间：
2008-03-01
影响因子：
3.2
通讯作者：
Kondo, Takao
中科院分区：
文献类型：
--
作者：
Murayama, Yoriko;Oyama, Tokitaka;Kondo, Takao
三种生物钟蛋白KaiA、KaiB和KaiC被认为是蓝细菌昼夜节律振荡器的重要组成部分，基于KaiB的化学振荡被认为是细长聚球藻PCC 7942的基本昼夜节律计时机制。定量研究了蓝藻细胞中kaiBC的转录和翻译，以阐明这些过程如何与化学振荡器偶联，使用的菌株中，昼夜节律振荡是在IPTG（异丙基-β-D-硫代半乳糖苷）的控制下。由IPTG触发的kaiBC启动子的阻遏动力学允许估计10小时的瞬时响应。该响应时间适合于蓝细菌转录和/或翻译以与基于Kai的振荡器匹配。有趣的是，kaiBC启动子活性和KaiC磷酸化显示出强大的昼夜节律，而trc启动子驱动的kaiBC mRNA水平和KaiC积累几乎是节律性的。这些结果表明，即使kaiBC表达是组成型的，也可以产生蓝藻昼夜节律。此外，在三种节律条件下，kaiBC启动子的激活与KaiC磷酸化比率的增加之间存在正相关性。基于这些观察，KaiC磷酸化比率可能是kaiBC启动子激活的主要因素。最后，我们定量地比较了磷酸化KaiC对kaiBC启动子的阻遏或去阻遏的阈值水平，发现该参数是阻遏kaiBC启动子的重要因素。
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.