蓝藻的生物钟,预测日常环境变化,通常包括由KaiA蛋白组成的标准振荡器,凯布,和KaiC。然而,几种蓝藻具有功能不明确的多种Kai蛋白同源物。特别是,集胞藻。PCC6803港口,除了一个典型的kaiABC基因簇(名为kaiAB1C1),另外两个kaiB和kaiC同源物(kaiB2、kaiB3、kaiC2、kaiC3)。这里,我们鉴定了一个嵌合KaiA同源物,命名为KaiA3,由位于kaiB3上游的基因编码。在N端,KaiA3类似于响应调节器接收器域,而其C端结构域类似于KaiA。同源性分析表明,几种蓝藻和其他细菌中存在KaiA3-KaiB3-KaiC3系统。使用集胞藻sp。PCC6803同源物,在KaiA3和KaiB3存在的情况下,我们观察到KaiC3磷酸化的昼夜节律振荡。kaiA3突变影响kaiC3磷酸化,导致兼养和化学异养条件下的生长缺陷。KaiC1和KaiC3表现出相位锁定的自由运行磷酸化节律。删除任一系统(取决于kaiAB1C1或取决于kaiA3B3C3)都会改变细胞反向散射节律的周期。此外,两个振荡器都需要保持高振幅,自持反向散射振荡,周期约为24小时,表明它们相互联系的性质。
The circadian clock of cyanobacteria, which predicts daily environmental changes, typically includes a standard oscillator consisting of proteins KaiA, KaiB, and KaiC. However, several cyanobacteria have diverse Kai protein homologs of unclear function. In particular,
Synechocystis sp. PCC 6803 harbours, in addition to a canonical kaiABC gene cluster (named kaiAB1C1), two further kaiB and kaiC homologs (kaiB2, kaiB3, kaiC2, kaiC3). Here, we identify a chimeric KaiA homolog, named KaiA3, encoded by a gene located upstream of kaiB3. At the N-terminus, KaiA3 is similar to response-regulator receiver domains, whereas its C-terminal domain resembles that of KaiA. Homology analysis shows that a KaiA3-KaiB3-KaiC3 system exists in several cyanobacteria and other bacteria. Using the
Synechocystis sp. PCC 6803 homologs, we observe circadian oscillations in KaiC3 phosphorylation in vitro in the presence of KaiA3 and KaiB3. Mutations of kaiA3 affect KaiC3 phosphorylation, leading to growth defects under both mixotrophic and chemoheterotrophic conditions. KaiC1 and KaiC3 exhibit phase-locked free-running phosphorylation rhythms. Deletion of either system (∆kaiAB1C1 or ∆kaiA3B3C3) alters the period of the cellular backscattering rhythm. Furthermore, both oscillators are required to maintain high-amplitude, self-sustained backscatter oscillations with a period of approximately 24 h, indicating their interconnected nature.