Control of mammalian circadian rhythm by CKIepsilon-regulated proteasome-mediated PER2 degradation.
Control of mammalian circadian rhythm by CKIepsilon-regulated proteasome-mediated PER2 degradation.
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发表时间：
2005
影响因子：
5.3
通讯作者：
E. Eide;Margaret F. Woolf;Heeseog Kang;P. Woolf;W. Hurst;F. Camacho;E. Vielhaber;A. Giovanni;D. Virshup
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文献类型：
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作者：
E. Eide;Margaret F. Woolf;Heeseog Kang;P. Woolf;W. Hurst;F. Camacho;E. Vielhaber;A. Giovanni;D. Virshup
哺乳动物昼夜节律调节蛋白PER 1和PER 2经历每日的积累循环，随后是磷酸化和降解。虽然这些抑制剂的磷酸化调节的蛋白水解被假定为是必不可少的时钟的功能，这一过程的抑制尚未被证明会改变哺乳动物的昼夜节律。我们已经开发了基于细胞的PER 2降解模型。由细胞渗透性蛋白磷酸酶抑制剂calyculin A诱导的小鼠PER 2（mPER 2）过度磷酸化，随后迅速被26 S蛋白酶体泛素化和降解。蛋白酶体介导的降解在生物钟中至关重要，因为蛋白酶体抑制剂导致Rat-1细胞的昼夜节律周期显著延长。已假定CKI 3（酪蛋白激酶I3）引发PER 2降解。支持这一观点，CKI 3抑制也导致同步化的Rat-1细胞中的昼夜节律周期显著延长。CK 1 β抑制还减缓细胞中PER 2的降解。CK ε介导的PER 2磷酸化将泛素连接酶接头蛋白β-TrCP募集到特定位点，显性负性β-TrCP阻断mPER 2的磷酸化依赖性降解。这些结果提供了一个生化机制和功能的相关性，所观察到的哺乳动物PER 2的磷酸化降解周期。基于细胞培养的生化分析结合基于细胞的节律测量补充遗传学研究，以阐明控制哺乳动物生物钟的基本机制。
The mammalian circadian regulatory proteins PER1 and PER2 undergo a daily cycle of accumulation followed by phosphorylation and degradation. Although phosphorylation-regulated proteolysis of these inhibitors is postulated to be essential for the function of the clock, inhibition of this process has not yet been shown to alter mammalian circadian rhythm. We have developed a cell-based model of PER2 degradation. Murine PER2 (mPER2) hyperphosphorylation induced by the cell-permeable protein phosphatase inhibitor calyculin A is rapidly followed by ubiquitination and degradation by the 26S proteasome. Proteasome-mediated degradation is critically important in the circadian clock, as proteasome inhibitors cause a significant lengthening of the circadian period in Rat-1 cells. CKIepsilon (casein kinase Iepsilon) has been postulated to prime PER2 for degradation. Supporting this idea, CKIepsilon inhibition also causes a significant lengthening of circadian period in synchronized Rat-1 cells. CKIepsilon inhibition also slows the degradation of PER2 in cells. CKIepsilon-mediated phosphorylation of PER2 recruits the ubiquitin ligase adapter protein beta-TrCP to a specific site, and dominant negative beta-TrCP blocks phosphorylation-dependent degradation of mPER2. These results provide a biochemical mechanism and functional relevance for the observed phosphorylation-degradation cycle of mammalian PER2. Cell culture-based biochemical assays combined with measurement of cell-based rhythm complement genetic studies to elucidate basic mechanisms controlling the mammalian clock.