Circadian SCN-Liver Axis in the Neuroendocrine Response to Calorie Restriction

在哺乳动物中，下丘脑的起搏器时钟同步外周组织的时钟以进行临时分区 氧化和还原代谢途径使燃料利用与养分供应相一致。然而，昼夜节律 大脑和外周组织的时钟将营养状态与转录结合起来，促进能量节约 在睡眠和营养匮乏的情况下，新陈代谢的动态平衡仍然不清楚。一条令人兴奋的线索 营养信号是如何控制昼夜节律转录的，这是我们团队和其他人发现的 烟酰胺腺嘌呤二核苷酸(NAD+)和依赖NAD+的脱乙酰酶SIRT1调节昼夜节律 通过核心时钟抑制因子PER2的翻译后修饰实现行为和线粒体节律， 这表明NAD+-SIRT1控制神经元和外周细胞的时钟周期。相互转换 在氧化还原反应中，NAD+及其还原形式NADH依赖于营养状态。在新的结果中 在我们的第一篇论文发表后，我们展示了在健康的卡路里限制期间NADH在肝脏中的积累。 抑制SIRT1，降低白天体温和氧化代谢。令人惊讶的是，减少NADH 短乳杆菌(LbNOX)通过肝脏转导水形成NADH氧化酶的水平解除抑制 SIRT1并增强新陈代谢和转录的氧化循环。此外，我们新生成的PER2K680Q 对SIRT1诱导的脱乙酰化具有抵抗力的拟乙酰敲击小鼠表现出深刻的周期 延长，而视交叉上核(SCN)的时钟消融则取消节律性摄食和 生热作用。我们现在准备利用创新的遗传工具和昼夜节律协议来剖析 生物钟促进睡眠期间的能量恒定，并在以下水平上适应卡路里限制 肝脏(目标1)和下丘脑起搏神经元(目标2)。目标1将专门测试营养素的假设 时钟检测涉及NAD(H)-SIRT1信令。我们建议剖析氧化还原态在时钟中的作用 通过使用基因控制NAD(H)水平来限制睡眠和卡路里的功能和代谢 LbNOx联合肝钟消融或PER2K680Q乙酰化敲击小鼠。Aim 2将检查 下丘脑起搏神经元亚型在同步产热、摄食和代谢中的作用 睡眠节律和对卡路里限制的适应性反应 CRISPR-CAS9时钟消融、功能失得性研究和基于投影的化学发生操作 起搏器神经元。总而言之，我们拟议的研究将阐明涉及以下方面的生理机制 在睡眠-唤醒周期中保持能量恒定，以及时钟适应如何有助于健康 低热量饮食的好处。

In mammals, the hypothalamic pacemaker clock synchronizes peripheral tissue clocks to temporally partition oxidative and reductive metabolic pathways to align fuel utilization with nutrient availability. Yet how the circadian clock in brain and peripheral tissues integrates nutrient state with transcription to promote energy conservation and metabolic homeostasis during sleep and in nutrient scarce conditions remains obscure. An exciting clue as to how nutrient signals control circadian transcription emerged from the discovery in our group and others that nicotinamide adenine dinucleotide (NAD+) and the NAD+-dependent deacetylase SIRT1 regulate circadian behavioral and mitochondrial rhythms through posttranslational modification of the core clock repressor PER2, indicating that NAD+-SIRT1 controls clock cycles within both neurons and peripheral cells. Interconversion of NAD+ with its reduced form NADH during redox reactions is dependent upon nutrient state. In new results published after our first submission, we show that NADH accumulation in liver during healthful calorie restriction inhibits SIRT1 and reduces daytime body temperature and oxidative metabolism. Surprisingly, reducing NADH levels through hepatic transduction of the water-forming NADH oxidase Lactobacillus brevis (LbNOX) disinhibits SIRT1 and augments oxidative cycles of metabolism and transcription. Further, our newly-generated PER2K680Q acetyl-mimetic knockin mice, which are resistant to SIRT1-induced deacetylation, exhibit profound period lengthening, while clock ablation in the suprachiasmatic nucleus (SCN) abrogates rhythmic feeding and thermogenesis. We are now poised with innovative genetic tools and circadian protocols to dissect how the circadian clock promotes energy constancy during sleep and in adaptation to calorie restriction at the level of the liver (Aim 1) and hypothalamic pacemaker neurons (Aim 2). Aim 1 will specifically test the hypothesis that nutrient sensing by the clock involves NAD(H)-SIRT1 signaling. We propose to dissect the role of redox state in clock function and metabolism during sleep and calorie restriction by genetically manipulating NAD(H) levels using LbNox in combination with hepatic clock ablation or PER2K680Q acetyl-mimetic knockin mice. Aim 2 will examine the role of hypothalamic pacemaker neuron subtypes in synchronizing thermogenesis, feeding, and metabolic rhythms with sleep and in the adaptive response to calorie restriction by utilizing an innovative combination of CRISPR-Cas9 clock ablation, loss and gain of function studies, and projection-based chemogenetic manipulation of pacemaker neurons. Collectively, our proposed studies will elucidate circadian mechanisms involved in maintenance of energy constancy across the sleep-wake cycle and how clock adaptations contribute to health benefits of hypocaloric diet.