The regulatory role of heme in circadian control

24小时生物钟是基本的内部计时系统，在几乎所有高等生物体（包括一些细菌）中产生行为，生理和代谢的每日节律。当生物钟被打乱时，如在长期轮班工作中，人类健康的恶化是显著的，据报道，抑郁症、睡眠障碍、心血管问题甚至癌症的水平升高。因此，了解生物钟在分子水平上的工作方式将对人类和动物健康产生重大益处，因为它将为设计新的治疗干预措施以改善这些负面副作用提供机会。在昆虫和脊椎动物中，生物钟依赖于两个关键的转录因子，CLOCK和BMAL 1，它们与生物钟靶基因的DNA结合并激活它们的表达。它们的作用至关重要，文献中出现了令人信服的证据，证明金属元素铁-以血红素铁的形式-与时钟结合。血红素是一种小的有机分子，中心含有铁，能够结合氧（O2）、一氧化碳（CO）和一氧化氮（NO）等配体。这些配体被认为是重要的分子信号，例如，生物体的代谢状态（通过这些配体浓度的变化）可能会向血红素发出信号，以改变下游时钟基因调节的方式改变时钟的特性。然而，血红素介导的时钟调节机制尚未建立。我们将探索血红素和气体结合到人类CLOCK转录因子，并确定血红素-CLOCK相互作用的结构。我们还将研究血红素如何影响CLOCK与其伴侣BMAL 1结合的方式，以及血红素对CLOCK-BMAL 1二聚体与其DNA靶序列的物理相互作用的重要性。最后，我们将使用苍蝇细胞研究血红素是如何参与的方式时钟BMAL 1刺激目标时钟基因的转录使用一个简单的发光读出。

The 24 hour circadian clock is the fundamental internal timekeeping system that generates daily rhythms of behaviour, physiology and metabolism in nearly all higher organisms, including some bacteria. When the clock is disrupted, as in chronic shift work, the deterioation of human health is significant, and elevated levels of depression, sleep disorders, cardiovascular problems and even cancer, have been reported. Consequently, understanding how the clock works at the molecular level will have significant benefits for human and animal health because it will provide opportunities to design novel therapeutic interventions to ameliorate these negative side-effects. In insects and vertebrates, the clock depends on two critical transcription factors, CLOCK and BMAL1 that bind to the DNA of clock target genes and activate their expression. Their role is crucial and there is convincing evidence emerging in the literature that the metallic element iron - in the form of heme iron - binds to CLOCK. Heme is a small, organic molecule with iron at the centre with an ability to bind ligands such as oxygen (O2), carbon monoxide (CO) and nitric oxide (NO). These ligands are recognised as important molecular signals so that, for example, the metabolic state of the organism (via changes in the concentrations of these ligands) might signal to heme, altering the properties of CLOCK in such a way that the regulation of downstream clock genes is changed. However, the mechanisms of heme-mediated regulation of the clock have yet to be established.We will explore heme and gas binding to the human CLOCK transcription factor and determine the structure for the heme-CLOCK interaction. We will also study how heme affects the way in which CLOCK binds to its partner BMAL1, and also how heme is important for the physical interaction of this CLOCK-BMAL1 dimer with its DNA target sequence. Finally, we shall use fly cells to study how heme is involved in the way CLOCK-BMAL1 stimulates transcription of target clock genes using a simple luminescence readout.