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Abstract Post-translational modifications (PTMs) couple feed-fast cycles to diurnal rhythms. However, it remains largely uncharacterized whether and how meal timing organizes diurnal rhythms beyond the transcriptome. Here, we systematically profile the daily rhythms of the proteome, four PTMs (phosphorylation, ubiquitylation, succinylation and N-glycosylation) and the lipidome in the liver from young female mice subjected to either day/sleep time-restricted feeding (DRF) or night/wake time-restricted feeding (NRF). We detect robust daily rhythms among different layers of omics with phosphorylation the most nutrient-responsive and succinylation the least. Integrative analyses reveal that clock regulation of fatty acid metabolism represents a key diurnal feature that is reset by meal timing, as indicated by the rhythmic phosphorylation of the circadian repressor PERIOD2 at Ser971 (PER2-pSer971). We confirm that PER2-pSer971 is activated by nutrient availability in vivo. Together, this dataset represents a comprehensive resource detailing the proteomic and lipidomic responses by the liver to alterations in meal timing.

Background: Meal timing resets circadian clocks in peripheral tissues, such as the liver, in seven days without affecting the phase of the central clock located in the suprachiasmatic nucleus (SCN) of the hypothalamus. Anterior hypothalamus plays an essential role in energy metabolism, circadian rhythm, and stress response. However, it remains to be elucidated whether and how anterior hypothalamus adapts its circadian rhythms to meal timing. Methods: Here, we applied transcriptomics to profile rhythmic transcripts in the anterior hypothalamus of nocturnal female mice subjected to day- (DRF) or night (NRF)-time restricted feeding for seven days. Results: This global profiling identified 128 and 3,518 rhythmic transcripts in DRF and NRF, respectively. NRF entrained diurnal rhythms among 990 biological processes, including ‘Electron transport chain’ and ‘Hippo signaling’ that reached peak time in the late sleep and late active phase, respectively. By contrast, DRF entrained only 20 rhythmic pathways, including ‘Cellular amino acid catabolic process’, all of which were restricted to the late active phase. The rhythmic transcripts found in both DRF and NRF tissues were largely resistant to phase entrainment by meal timing, which were matched to the action of the circadian clock. Remarkably, DRF for 36 days partially reversed the circadian clock compared to NRF. Conclusions: Collectively, our work generates a useful dataset to explore anterior hypothalamic circadian biology and sheds light on potential rhythmic processes influenced by meal timing in the brain (www.circametdb.org.cn).

High-fat diet (HFD) feeding rewires circadian rhythms of peripheral organs including the liver and adipose tissue. While the liver has been extensively studied, it remains largely unknown whether and how HFD organizes circadian biology in adipose tissue. Here, we took a systems approach to profile the diurnal transcriptome of adipose tissue in diet-induced obese mice either fed a low-fat diet (LFD) that reduces weight or still fed HFD. We detected about 200 and 2,500 diurnal genes in HFD and LFD, respectively. Pathway analysis revealed that rhythmic pathways in HFD are represented by circadian rhythm, ribosome biogenesis, and nucleosome organization, whereas those in LFD are represented by myeloid cell function. Remarkably, the majority of the circadian clock genes, except Clock, exhibited robust diurnal rhythm in the adipose tissue of HFD-fed mice. Analysis of mRNAs and proteins in another cohort of HFD-fed mice confirmed that Clock lost rhythmicity at the transcript, but not protein level. Diet reversal to LFD specifically restored diurnal difference of the Clock transcripts in adipose tissue. We matched transcriptomics data with global profiling of neutral lipids and found that lipid metabolism catalyzed by triglycerol hydrolase Ces1d is a key circadian feature that is activated by diet reversal. Together, our work defines the circadian signatures in the adipose tissue of diet-induced obese mice, and their flexibility upon dietary intervention, thereby shedding light on potential clock-modulated tissue-specific pathways during obesity.

Summary: Inverted feeding is a paradigm to study synchronization of circadian clocks by feeding rhythm in tissues more directly. Here, we provide a protocol for performing inverted feeding in mice and analyzing circadian rhythmicity in mouse tissues. We describe setting up inverted feeding and performing tissue dissection, followed by RNA extraction and gene expression analysis, and lastly R software-based analysis of circadian rhythmicity. This protocol can be combined with the use of CircaMetDB database for mechanistic studies of inverted feeding.For complete details on the use and execution of this protocol, please refer to Xin et al. (2021).

The circadian clock coordinates physiology, metabolism, and behavior with the 24-h cycles of environmental light. Fundamental mechanisms of how the circadian clock regulates organ physiology and metabolism have been elucidated at a rapid speed in the past two decades. Here we review circadian networks in more than six organ systems associated with complex disease, which cluster around metabolic disorders, and seek to propose critical regulatory molecules controlled by the circadian clock (named clock-controlled checkpoints) in the pathogenesis of complex disease. These include clock-controlled checkpoints such as circadian nuclear receptors in liver and muscle tissues, chemokines and adhesion molecules in the vasculature. Although the progress is encouraging, many gaps in the mechanisms remain unaddressed. Future studies should focus on devising time-dependent strategies for drug delivery and engagement in well-characterized organs such as the liver, and elucidating fundamental circadian biology in so far less characterized organ systems, including the heart, blood, peripheral neurons, and reproductive systems.

Post-translational O-linked β-N acetylglucosamine (O-GlcNAc) modification acts as a nutrient-sensing mechanism. Here the authors report that O-GlcNAc transferase inhibits adipose tissue lipolysis via O-GlcNAcylation of the lipid droplet protein perilipin 1 and thus promotes diet-induced obesity.

Summary: Time of eating synchronizes circadian rhythms of metabolism and physiology. Inverted feeding can uncouple peripheral circadian clocks from the central clock located in the suprachiasmatic nucleus. However, system-wide changes of circadian metabolism and physiology entrained to inverted feeding in peripheral tissues remain largely unexplored. Here, we performed a 24-h global profiling of transcripts and metabolites in mouse peripheral tissues to study the transition kinetics during inverted feeding, and revealed distinct kinetics in phase entrainment of diurnal transcriptomes by inverted feeding, which graded from fat tissue (near-completely entrained), liver, kidney, to heart. Phase kinetics of tissue clocks tracked with those of transcriptomes and were gated by light-related cues. Integrated analysis of transcripts and metabolites demonstrated that fatty acid oxidation entrained completely to inverted feeding in heart despite the slow kinetics/resistance of the heart clock to entrainment by feeding. This multi-omics resource defines circadian signatures of inverted feeding in peripheral tissues (www.CircaMetDB.org.cn).

Endocannabinoid signaling regulates food intake and is a potential therapeutic target for obesity. Here the authors show that adipocyte O-GlcNAc transferase (OGT) is required for high fat diet-induced hyperphagia via transcriptional activation of de novo lipid desaturation and accumulation of an endogenous appetite-inducing cannabinoid.

Summary: Deterioration of metabolic health is a hallmark of aging and generally assumed to be detrimental to longevity. Exposure to a high-calorie diet impairs metabolism and accelerates aging; conversely, calorie restriction (CR) prevents age-related metabolic diseases and extends lifespan. However, it is unclear whether preservation of metabolic health is sufficient to extend lifespan. We utilized a genetic mouse model lacking Fabp4/5 that confers protection against metabolic diseases and shares molecular and lipidomic features with CR to address this question. Fabp-deficient mice exhibit extended metabolic healthspan, with protection against insulin resistance and glucose intolerance, inflammation, deterioration of adipose tissue integrity, and fatty liver disease. Surprisingly, however, Fabp-deficient mice did not exhibit any extension of lifespan. These data indicate that extension of metabolic healthspan in the absence of CR can be uncoupled from lifespan, indicating the potential for independent drivers of these pathways, at least in laboratory mice. : Deterioration of metabolic health is a hallmark of aging and generally thought to be detrimental to longevity. Charles et al. utilize FABP-deficient mice as a model to demonstrate that the preservation of metabolic health in this model persists throughout life, even under metabolic stress, but does not increase longevity. Keywords: fatty acid binding protein, aging, calorie restriction, metabolic health, inflammation, metaflammation, diabetes, obesity, de novo lipogenesis

Members of the nuclear receptor superfamily have vital roles in regulating immunity and inflammation. The founding member, glucocorticoid receptor (GR), is the prototype to demonstrate immunomodulation via transrepression of the AP-1 and NF-κB signaling pathways. Peroxisome proliferator-activated receptors (PPARs) have emerged as key regulators of inflammation. This review examines the history and current advances in nuclear receptor regulation of inflammation by the crosstalk with AP-1 and NF-κB signaling, focusing on the roles of GR and PPARs. A better understanding of the molecular mechanism by which nuclear receptors inhibit proinflammatory signaling pathways will enable novel therapies to treat chronic inflammation.

RIG-I and MDA5 are cytoplasmic sensors that recognize different species of viral RNAs, leads to activation of the transcription factors IRF3 and NF-kappaB, which collaborate to induce type I interferons. In this study, we identified REUL, a RING-finger protein, as a specific RIG-I-interacting protein. REUL was associated with RIG-I, but not MDA5, through its PRY and SPRY domains. Overexpression of REUL potently potentiated RIG-I-, but not MDA5-mediated downstream signalling and antiviral activity. In contrast, the RING domain deletion mutant of REUL suppressed Sendai virus (SV)-induced, but not cytoplasmic polyI:C-induced activation of IFN-beta promoter. Knockdown of endogenous REUL by RNAi inhibited SV-triggered IFN-beta expression, and also increased VSV replication. Full-length RIG-I, but not the CARD domain deletion mutant of RIG-I, underwent ubiquitination induced by REUL. The Lys 154, 164, and 172 residues of the RIG-I CARD domain were critical for efficient REUL-mediated ubiquitination, as well as the ability of RIG-I to induce activation of IFN-beta promoter. These findings suggest that REUL is an E3 ubiquitin ligase of RIG-I and specifically stimulates RIG-I-mediated innate antiviral activity.

Recruitment of O-GlcNAc transferase (OGT) to promoters plays an important role in gene repression. Glucocorticoid signaling represses the transcriptional activities of NF-?B and AP-1 through direct binding, yet the molecular mechanisms remain to be elucidated. Here we report that OGT is an important component of GR-mediated transrepression. OGT associates with ligand-bound GR in a multi-protein repression complex. Overexpression of OGT potentiates the GR transrepression pathway, whereas depletion of endogenous OGT by RNA interference abolishes the repression. The recruitment of OGT by GR leads to increased O-GlcNAcylation and decreased phosphorylation of RNA polymerase II on target genes. Functionally, overexpression of OGT enhances glucocorticoid-induced apoptosis in resistant cell lines while knockdown of OGT prevents sensitive cell lines from apoptosis. These studies identify a molecular mechanism of GR transrepression, and highlight the function of O-GlcNAc in hormone signaling. [ABSTRACT FROM AUTHOR]

Many intracellular proteins are reversibly modified by Olinked GlcNAc (O-GlcNAc), a post-translational modification that dynamically regulates fundamental cellular processes in response to diverse environmental cues. Accumulating evidence indicates that both excess and deficiency of protein O-GlcNAcylation can have deleterious effects on the cell, suggesting that maintenance of O-GlcNAc homeostasis is essential for proper cellular function. However, the mechanisms through which O-GlcNAc homeostasis is maintained in the physiologic state and altered in the disease state have not yet been investigated. Here, we demonstrate the existence of a homeostaticmechanisminvolving mutual regulation of the O-GlcNAc-cycling enzymes O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) at the transcriptional level. Specifically, we found that OGA promotes Ogt transcription through cooperation with the histone acetyltransferase p300 and transcription factor CCAAT/enhancer-binding protein β (C/EBPβ). To examine the role of mutual regulation of OGT and OGA in the disease state, we analyzed gene expression data from human cancer data sets, which revealed that OGT and OGA expression levels are highly correlated in numerous human cancers, particularly in pancreatic adenocarcinoma. Using a KrasG12D-driven primary mouse pancreatic ductal adenocarcinoma (PDAC) cell line, we found that inhibition of extracellular signal-regulated kinase (ERK) signaling decreases OGA glycosidase activity and reduces OGT mRNA and protein levels, suggesting that ERK signaling may alter O-GlcNAc homeostasis in PDAC by modulating OGA-mediated Ogt transcription. Our study elucidates a transcriptional mechanism that regulates cellularO-GlcNAc homeostasis, which may lay a foundation for exploring O-GlcNAc signaling as a therapeutic target for human disease. [ABSTRACT FROM AUTHOR]