Sun- versus moonlight responses and molecular interactions of a marine Cryptochrome

海洋隐花色素的太阳光与月光反应以及分子相互作用

• 批准号: 538823817
• 负责人: Professorin Dr. Eva Wolf
• 金额: --
• 依托单位: Arbeitsgruppe Strukturelle Chronobiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/538823817?language=en
• 关键词: Sun; versus; moonlight; responses; molecular

Many marine organisms time their maturation and reproduction according to the lunar cycle using an inner monthly calendar, also referred to as circalunar clock or circalunar oscillator. The marine bristle worm Platynereis dumerilii (Pdu) has a light-sensitive cryptochrome (Cry) protein, called L-Cry, that can discriminate between moonlight and sunlight and between different moon phases. This enables the worm to synchronize its reproduction to the full moon phase via its circalunar clock. We previously showed that L-Cry can form three distinct molecular states with unique biochemical and functional properties: 1. a dimeric, predominantly nuclear, dark state with its light-sensing cofactor flavine adenine dinucleotide (FAD) in the fully oxidized form, 2. a monomeric, predominantly cytosolic, photoactivated sunlight state with a fully reduced anionic FAD°- radical and 3. a half-reduced, predominantly nuclear, moonlight state. Here, we will further elucidate the molecular mechanisms underlying the differential responses of L-Cry to sunlight and moonlight, using a combination of protein biochemical, spectroscopic, 3D-structural and cell-based approaches. Our specific aims are to understand the molecular basis of the exceptionally high light sensitivity of L-Cry that enables it to sense very dim moonlight, to uncover the molecular structure of the enigmatic L-Cry moonlight state, and to dissect conformational rearrangements and intermediate states occurring during the transitions between the dark-, sunlight- and moonlight states of L-Cry. Furthermore, we will determine the upper and lower limits of the light intensity ranges, that lead to the formation of functional moonlight- or sunlight states of L-Cry. This will enable us to predict the impact of anthropogenic artificial nocturnal light pollution on L-Cry moonlight signalling and on the circalunar behaviour of the worms. To elucidate the hitherto unknown molecular basis of circalunar oscillators, we will also analyse L-Cry’s downstream signalling to the circalunar clock. To this end, we will search for new L-Cry interactors using pulldown/mass spectrometry, and mechanistically and functionally characterize their interactions with L-Cry in vitro and in vivo.

许多海洋生物使用内部每月日历（也称为圆形时钟或圆形振荡器）根据月球周期的成熟和繁殖时间。海洋猪猪蠕虫铂雷氏（Platyeris dumerilii）（PDU）具有一种光敏的加密色素（Cry）蛋白，称为L-Cry，可以区分月光和阳光，以及在不同的月相之间。这使蠕虫能够通过其圆形时钟将其繁殖与满月相同步。我们先前表明，L-cry可以形成具有独特生化和功能特性的三个不同的分子状态：1。二聚体，主要是核的，黑暗状态，其光感应辅助剂黄酮丁胺二核苷酸（FAD），完全氧化的形式，2。一种。半还原，主要是核月光状态。在这里，我们将进一步阐明L-Cry对阳光和月光差异反应的分子机制，结合使用蛋白质生化，光谱，3D结构和基于细胞的方法。我们的具体目的是了解L-cry的异常高光灵敏度的分子基础，使其能够感知到非常昏暗的月光，发现神秘的L-Cry月光状态的分子结构，并解剖构象的重排，并在L-CRY的黑暗，太阳光和月光状态之间发生过渡期间发生的中间状态。此外，我们将确定光强度范围的上和下限，从而导致L-Cry的功能性月光或阳光状态的形成。这将使我们能够预测人为的人工夜间光污染对L-Cry Moonlight信号传导以及蠕虫圆形行为的影响。为了阐明圆形振荡器的隐藏到未知的分子基础，我们还将分析L-Cry的下游信号传导到圆形时钟。为此，我们将使用下拉/质量光谱法搜索新的L-Cry交互剂，并在机械上和功能上表征其与L-Cry在体外和体内的相互作用。