Institut de Neurobiologie Alfred Fessard, Centre National de la Recherche Scientifique Unité Propre de Recherche 3294, Gif-sur-Yvette, France ; Département de Biologie, Université Paris Sud, Orsay, France.
PLoS Biol. 2013;11(8):e1001645. doi: 10.1371/journal.pbio.1001645. Epub 2013 Aug 27.
Phosphorylation is a pivotal regulatory mechanism for protein stability and activity in circadian clocks regardless of their evolutionary origin. It determines the speed and strength of molecular oscillations by acting on transcriptional activators and their repressors, which form negative feedback loops. In Drosophila, the CK2 kinase phosphorylates and destabilizes the PERIOD (PER) and TIMELESS (TIM) proteins, which inhibit CLOCK (CLK) transcriptional activity. Here we show that CK2 also targets the CLK activator directly. Downregulating the activity of the catalytic α subunit of CK2 induces CLK degradation, even in the absence of PER and TIM. Unexpectedly, the regulatory β subunit of the CK2 holoenzyme is not required for the regulation of CLK stability. In addition, downregulation of CK2α activity decreases CLK phosphorylation and increases per and tim transcription. These results indicate that CK2 inhibits CLK degradation while reducing its activity. Since the CK1 kinase promotes CLK degradation, we suggest that CLK stability and transcriptional activity result from counteracting effects of CK1 and CK2.
磷酸化是生物钟中蛋白质稳定性和活性的关键调节机制，无论其进化起源如何。它通过作用于转录激活因子及其阻遏物来决定分子振荡的速度和强度，这些因子形成负反馈回路。在果蝇中，CK2 激酶磷酸化并使 PERIOD（PER）和 TIMELESS（TIM）蛋白失稳，从而抑制 CLOCK（CLK）转录活性。在这里，我们表明 CK2 还直接靶向 CLK 激活剂。下调 CK2 催化α亚基的活性会诱导 CLK 降解，即使在没有 PER 和 TIM 的情况下也是如此。出乎意料的是，CK2 全酶的调节β亚基对于 CLK 稳定性的调节不是必需的。此外，下调 CK2α活性会降低 CLK 的磷酸化水平，增加 per 和 tim 的转录。这些结果表明，CK2 在降低 CLK 活性的同时抑制其降解。由于 CK1 激酶促进 CLK 降解，我们认为 CLK 的稳定性和转录活性是由 CK1 和 CK2 的拮抗作用产生的。
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