The pentose phosphate pathway regulates the circadian clock

The circadian clock is a ubiquitous timekeeping system that organizes the behavior and physiology of organisms over the day and night. The clockwork orchestrates a multitude of metabolic processes as illustrated by previous global transcriptomics and proteomics studies, and the existence of daily rhythms of reduction and oxidation (redox) in a range of diverse species. However, the reciprocal question of whether metabolism can alter the clockwork remains largely unaddressed. Here we identify the pentose phosphate pathway (PPP), a critical source of cellular reducing power in the form of NADPH, as an important modulator of circadian oscillations. We show that genetic and pharmacologic inhibition of the PPP perturbs circadian gene expression and metabolic rhythms in human cells. Pharmacologic inhibition of the PPP caused similar effects in mouse tissues, and altered the pattern of rhythmic behavior in Drosophila. These manipulations also altered genome wide DNA-binding activity of the circadian transcription factors BMAL1 and CLOCK through a mechanism likely involving the redox-sensitive histone acetyltransferase p300. Thus, disruption of the PPP regulates circadian rhythms via modulation of NADPH metabolism, highlighting redox reactions as a novel connector of metabolic cycles and transcriptional oscillations in nucleated cells. Overall design: RNA-Seq time-course experiment in U2OS cells treated with 6AN or control over 48 hours at 4h resolution. ChIP-Seq of BMAL1, CLOCK, H3K4me3 and H3K9ac in U2OS cells treated with 6AN or control for 24h.

昼夜节律钟（circadian clock）是一种普遍存在的计时系统，可在昼夜周期中调控生物体的行为与生理活动。过往的全局转录组学、蛋白质组学研究，以及多种物种中普遍存在的每日还原-氧化（redox）节律现象，均证实该生物钟调控着大量代谢过程。然而，代谢是否能够反过来调控生物钟这一反向问题，目前仍未得到充分解答。本研究鉴定出磷酸戊糖途径（pentose phosphate pathway, PPP）——以烟酰胺腺嘌呤二核苷酸磷酸（NADPH）形式提供细胞还原力的关键通路——是昼夜节律振荡的重要调控因子。研究发现，对PPP进行遗传与药理学抑制，会扰乱人类细胞中的昼夜节律基因表达与代谢节律。对PPP进行药理学抑制，在小鼠组织中也可产生类似效应，并改变果蝇的节律行为模式。这类干预手段还会通过一种可能涉及氧化还原敏感型组蛋白乙酰转移酶p300的机制，改变昼夜节律转录因子BMAL1与CLOCK的全基因组DNA结合活性。综上，PPP的失衡可通过调控NADPH代谢来调节昼夜节律，这表明还原氧化反应是有核细胞中代谢周期与转录振荡之间的新型连接通路。实验设计：在经6AN或对照试剂处理的U2OS细胞中开展48小时、分辨率为4小时的RNA测序（RNA-Seq）时间序列实验；在经6AN或对照试剂处理的U2OS细胞中开展时长24小时的BMAL1、CLOCK、H3K4me3及H3K9ac染色质免疫沉淀测序（ChIP-Seq）实验。