CULLIN-3 controls TIMELESS oscillations in the Drosophila circadian clock.
CULLIN-3 controls TIMELESS oscillations in the Drosophila circadian clock.
复制标题
DOI：
10.1371/journal.pbio.1001367
复制
发表时间：
2012
期刊：
影响因子：
9.8
通讯作者：
Rouyer F
中科院分区：
文献类型：
--
作者：
Grima B;Dognon A;Lamouroux A;Chélot E;Rouyer F
泛素连接酶CUL-3和SLMB通过控制TIMELESS振荡协同调节果蝇昼夜节律钟。真核生物的生物钟依赖于转录反馈回路。在果蝇中，PERIOD（PER）和TIMELESS（TIM）蛋白在夜间积累，抑制CLOCK（CLK）/CYCLE（CYC）转录复合物的活性，并在清晨被降解。PER和TIM振荡的控制在很大程度上取决于翻译后机制。它们涉及依赖于时钟蛋白的磷酸化、泛素化和蛋白酶体降解的光依赖性和光非依赖性途径。SLMB是基于CULLIN-1的E3泛素连接酶复合物的一部分，是磷酸化PER的昼夜降解所需的。我们在这里表明，CULLIN-3（CUL-3）是必需的PER和TIM振荡的昼夜节律控制。时钟神经元中Cul-3 RNA干扰或Cul-3显性负性形式的表达改变了运动行为并抑制了光暗周期中的PER和TIM振荡。在恒定的条件下，CUL-3失调诱导行为不稳定性，并迅速消除TIM循环，对PER的影响较慢。CUL-3在PER不存在的情况下更强烈地影响TIM积累，并与低磷酸化TIM形成蛋白质复合物。相比之下，SLMB在PER的存在下更强烈地影响TIM，并且优先与磷酸化TIM相关联。CUL-3和SLMB对TIM和PER具有相加作用，表明两种泛素化复合物在PER和TIM循环中具有不同的作用。因此，这项工作表明，CUL-3是果蝇时钟的一个新的组成部分，它在TIM振荡的控制中起着重要的作用。生物钟将生物体的生理和行为调整为日/夜周期，并依赖于产生每日转录振荡的分子反馈回路。在果蝇中，PERIOD（PER）和TIMELESS（TIM）蛋白协调时钟-它们在夜间积累，形成复合物，并在清晨抑制自己的基因表达。这种振荡的时间控制涉及PER和TIM蛋白的磷酸化、泛素化和蛋白酶体降解。SUPERNUMERARY LIMBS（SLMB）泛素连接酶在调控磷酸化PER和TIM降解中起关键作用。在这项研究中，我们研究了另一种泛素连接酶，CULLIN-3（CUL-3）的作用。我们发现，抑制CUL-3的活性，结果在苍蝇的休息/活动节律的废除和消除PER和TIM振荡。CUL-3在PER不存在的情况下与TIM的低磷酸化形式物理相互作用并形成复合物，从而允许其在夜间积累。相反，当PER存在时，SLMB优先与磷酸化TIM相互作用，有利于其降解。结果表明，CUL-3和SLMB分担工作，以控制PER和TIM蛋白在白天/黑夜周期的振荡。
The ubiquitin ligases CUL-3 and SLMB collaborate to regulate the Drosophila circadian clock by controlling TIMELESS oscillations. Eukaryotic circadian clocks rely on transcriptional feedback loops. In Drosophila, the PERIOD (PER) and TIMELESS (TIM) proteins accumulate during the night, inhibit the activity of the CLOCK (CLK)/CYCLE (CYC) transcriptional complex, and are degraded in the early morning. The control of PER and TIM oscillations largely depends on post-translational mechanisms. They involve both light-dependent and light-independent pathways that rely on the phosphorylation, ubiquitination, and proteasomal degradation of the clock proteins. SLMB, which is part of a CULLIN-1-based E3 ubiquitin ligase complex, is required for the circadian degradation of phosphorylated PER. We show here that CULLIN-3 (CUL-3) is required for the circadian control of PER and TIM oscillations. Expression of either Cul-3 RNAi or dominant negative forms of CUL-3 in the clock neurons alters locomotor behavior and dampens PER and TIM oscillations in light-dark cycles. In constant conditions, CUL-3 deregulation induces behavioral arrhythmicity and rapidly abolishes TIM cycling, with slower effects on PER. CUL-3 affects TIM accumulation more strongly in the absence of PER and forms protein complexes with hypo-phosphorylated TIM. In contrast, SLMB affects TIM more strongly in the presence of PER and preferentially associates with phosphorylated TIM. CUL-3 and SLMB show additive effects on TIM and PER, suggesting different roles for the two ubiquitination complexes on PER and TIM cycling. This work thus shows that CUL-3 is a new component of the Drosophila clock, which plays an important role in the control of TIM oscillations. Circadian clocks adjust the physiology and behavior of organisms to the day/night cycle and rely on molecular feedback loops that generate daily oscillations of transcription. In the Drosophila fruit fly, the PERIOD (PER) and TIMELESS (TIM) proteins coordinate the clock–they accumulate during the night, form a complex, and repress their own gene expression in the early morning. The temporal control of this oscillation involves the phosphorylation, ubiquitination, and proteasomal degradation of the PER and TIM proteins. The SUPERNUMERARY LIMBS (SLMB) ubiquitin ligase is known to play a key role in controlling the degradation of phosphorylated PER and TIM. In this study we investigated the role of another ubiquitin ligase, CULLIN-3 (CUL-3). We found that inhibition of CUL-3 activity results in the abolition of rest/activity rhythms in flies and flattens the PER and TIM oscillations. CUL-3 physically interacts and forms a complex with a lowphosphorylated version of TIM in the absence of PER, thereby allowing its accumulation during the night. In contrast, when PER is present SLMB preferentially interacts with phosphorylated TIM, favoring its degradation. The results suggest that CUL-3 and SLMB share the work to control the oscillations of the PER and TIM proteins during the day/night cycle.
登录
查看更多内容
影响因子：
64.5
作者：
Chiu JC;Ko HW;Edery I
通讯作者：
Edery I
影响因子：
5.3
作者：
Harrisingh, Marie C.;Wu, Ying;Nitabach, Michael N.
通讯作者：
Nitabach, Michael N.
影响因子：
64.8
作者：
Grima, B;Chèlot, E;Rouyer, F
通讯作者：
Rouyer, F
影响因子：
1.3
作者：
Klarsfeld, A;Leloup, JC;Rouyer, F
通讯作者：
Rouyer, F
影响因子：
4.6
作者：
Kent, D;Bush, EW;Hooper, JE
通讯作者：
Hooper, JE