BMAL1-dependent circadian oscillation of nuclear CLOCK: posttranslational events induced bv dimerization of transcriptional activators of the mammalian clock system
BMAL1-dependent circadian oscillation of nuclear CLOCK: posttranslational events induced bv dimerization of transcriptional activators of the mammalian clock system
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DOI：
10.1101/gad.1099503
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发表时间：
2003-08-01
影响因子：
10.5
通讯作者：
Antoch, MP
中科院分区：
文献类型：
--
作者：
Kondratov, RV;Chernov, MV;Antoch, MP
哺乳动物CLOCK和BMAL1是bHLH-PAS转录因子家族中的两个成员，它们代表着昼夜自我调节反馈环的正向元件。以异源二聚体的形式，它们驱动反应基因启动子中E-box增强子元件的转录。我们研究了内源性和异位表达的Clock和BMAL1蛋白的丰度、翻译后修饰、细胞定位。Clock的胞核/胞浆分布受昼夜节律调节。对携带不同胚系昼夜节律突变的小鼠胚胎成纤维细胞CLOCK的亚细胞定位分析表明，CLOCK核积累的昼夜节律依赖于BMAL1。Clock和BMAL1蛋白异位共表达后形成Clock/BMAL1复合体，然后是相互依赖的磷酸化，与Clock核转位和降解紧密相连。这种结合依赖的协同调节是CLOCK/BMAL1相互作用所特有的，因为没有其他PAS结构域蛋白能够与CLOCK或BMAL1形成复合体而产生类似的作用。重要的是，我们研究中描述的所有翻译后事件都伴随着活跃的反式激活复合体的形成，这证明了它们具有重要的功能作用。总之，这些结果为额外水平的昼夜节律系统控制提供了证据，该控制基于对转录活性的调节或/和时钟/BMAL1复合体的可用性。
Mammalian CLOCK and BMAL1 are two members of bHLH-PAS-containing family of transcription factors that represent the positive elements of circadian autoregulatory feedback loop. In the form of a heterodimer, they drive transcription from E-box enhancer elements in the promoters of responsive genes. We have examined abundance, posttranslational modifications, cellular localization of endogenous and ectopically expressed CLOCK and BMAL1 proteins. Nuclear/cytoplasm distribution of CLOCK was found to be under circadian regulation. Analysis of subcellular localization of CLOCK in embryo fibroblasts of mice carrying different germ-line circadian mutations showed that circadian regulation of nuclear accumulation of CLOCK is BMAL1-dependent. Formation of CLOCK/BMAL1 complex following ectopic coexpression of both proteins is followed by their codependent phosphorylation, which is tightly coupled to CLOCK nuclear translocation and degradation. This binding-dependent coregulation is specific for CLOCK/BMAL1 interaction, as no other PAS domain protein that can form a complex with either CLOCK or BMAL1 was able to induce similar effects. Importantly, all posttranslational events described in our study are coupled with active transactivation complex formation, which argues for their significant functional role. Altogether, these results provide evidence for an additional level of circadian system control, which is based on regulation of transcriptional activity or/and availability of CLOCK/BMAL1 complex.