Mechanism of Photosignaling by Drosophila Cryptochrome
Mechanism of Photosignaling by Drosophila Cryptochrome
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果蝇隐花色素的光信号传导机制
DOI：
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发表时间：
2013
影响因子：
4.8
通讯作者：
A. Sancar
中科院分区：
文献类型：
--
作者：
Nuri Ozturk;C. Selby;D. Zhong;A. Sancar
背景：尚不清楚隐花色素的氧化还原状态是否影响生物钟重置。结果：阻断隐花色素光还原不会损害隐花色素介导的永恒降解，并且光可以诱导隐花色素与氧化黄素的构象变化。结论：隐花色素可以氧化或还原形式介导其功能。意义：这项研究为隐花色素光信号传递提供了新的见解。隐花色素（CRY）是果蝇中主要的昼夜节律光感受器。吸收光后，dCRY 会发生构象变化，使其能够与 Timeless (dTIM) 以及分别泛素化 dTIM 和 dCRY 的两种不同的 E3 连接酶结合，从而导致其蛋白水解并重置昼夜节律的相位。纯化的 dCRY 含有氧化黄素 (FADox)，它很容易通过一组 3 个高度保守的色氨酸残基（色氨酸三联体）光还原为阴离子半醌。 dCRY 的晶体结构揭示了第四个色氨酸 (Trp-536) 作为潜在的电子供体。此前，我们报道了色氨酸三联体在果蝇细胞中光诱导的 dCRY 蛋白水解中没有发挥作用。在这里，我们研究了 Trp 三联体和 Trp-536 的作用，以及黄素的氧化还原状态对光诱导的 dCRY 和 dTIM 蛋白水解以及时钟重置的影响。我们发现，dCRY 的氧化 (FADox) 和还原 (FAD⨪) 形式在体外都会经历光诱导的构象变化，使 dCRY 能够结合 JET，并且通过果蝇中 dCRY 和 dTIM 的光诱导蛋白水解测量，阻断已知或假定的蛋白内电子转移反应的 Trp 三联体和 Trp-536 突变不会影响体内亮光或暗光下的 dCRY 光转导S2R+ 细胞。我们得出结论，dCRY 的氧化形式和还原形式都能够产生光信号。
Background: It is not known whether the redox status of cryptochrome affects circadian clock resetting. Results: Blocking cryptochrome photoreduction does not impair cryptochrome-mediated Timeless degradation and light can induce a conformational change in cryptochrome with oxidized flavin. Conclusion: Cryptochrome can mediate its function in either oxidized or reduced form. Significance: This study provides novel insight into cryptochrome photosignaling. Cryptochrome (CRY) is the primary circadian photoreceptor in Drosophila. Upon light absorption, dCRY undergoes a conformational change that enables it to bind to Timeless (dTIM), as well as to two different E3 ligases that ubiquitylate dTIM and dCRY, respectively, resulting in their proteolysis and resetting the phase of the circadian rhythm. Purified dCRY contains oxidized flavin (FADox), which is readily photoreduced to the anionic semiquinone through a set of 3 highly conserved Trp residues (Trp triad). The crystal structure of dCRY has revealed a fourth Trp (Trp-536) as a potential electron donor. Previously, we reported that the Trp triad played no role in photoinduced proteolysis of dCRY in Drosophila cells. Here we investigated the role of the Trp triad and Trp-536, and the redox status of the flavin on light-induced proteolysis of both dCRY and dTIM and resetting of the clock. We found that both oxidized (FADox) and reduced (FAD⨪) forms of dCRY undergo light-induced conformational change in vitro that enable dCRY to bind JET and that Trp triad and Trp-536 mutations that block known or presumed intraprotein electron transfer reactions do not affect dCRY phototransduction under bright or dim light in vivo as measured by light-induced proteolysis of dCRY and dTIM in Drosophila S2R+ cells. We conclude that both oxidized and reduced forms of dCRY are capable of photosignaling.
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