Nature Metabolism

Dietary restriction (DR) protects against metabolic disease, extends lifespan, and is associated with remodeling of tissue reactive oxygen species (ROS). ROS control biological adaptation through reversible oxidation of protein cysteines, yet the targets of DR-initiated redox signaling are unknown. Here we generate OxiDR, a tissue-resolved atlas of the cysteine redox proteome that quantifies oxidation state under DR. Rather than oxidizing the proteome broadly, DR selectively targets a high-amplitude set of cysteines in a tissue-specific manner, allowing systematic classification of biological processes subject to DR-mediated redox regulation. Among the cysteines most highly oxidized upon DR is Cys19 of the core autophagy protein ATG5. We show oxidation of Cys19 is required for ATG5-mediated autophagosome formation and for autophagy triggered by nutrient restriction in human cells and mice. Reversible oxidation of this cysteine promotes ATG5 binding to ATG10, thus forming the ATG5-ATG12 conjugate that lipidates LC3B/ATG8 and matures the autophagosome. In mice, loss of this redox switch prevents effective initiation of autophagy upon nutrient restriction, resulting in gross tissue pathology and rapid onset of mortality. The autophagic response to nutrient restriction is thus gated by oxidation of a single cysteine.

Aging populations face a growing burden of neurodegenerative disease and behavioral decline. The microbiota is increasingly implicated in host physiology, highlighting an urgent need to understand how gut microbes modulate brain function and late life health. Here, we develop a high-throughput microbe-host screening platform to systematically dissect how bacterial genes shape organismal behavior. Using the Escherichia coli Keio library of 3,985 distinct non-essential single-gene deletion mutants in combination with Caenorhabditis elegans, we quantitatively profiled 768 behavioral features per bacterial clone and uncovered a coherent module of bacterial genes involved in iron homeostasis that robustly reprogrammed host behavior. Bacterial omics analyses revealed that alterations in iron homeostasis amplify siderophore production in a TCA cycle-dependent manner, and loss of enterobactin biosynthesis abolished the bacteria-induced changes in worm movement. Transcriptional, molecular and functional studies demonstrated that bacterial enterobactin activates host neuromuscular responses to shape both lifespan and healthspan in a manner dependent on the mitochondrial unfolded protein response. Thermal proteome profiling identified mitochondrial glutaryl-CoA dehydrogenase and the electron-transfer flavoproteins A and B as enterobactin-binding targets that control mitochondrial function. Finally, genetic manipulation of neuronal mitochondrial and cytosolic reactive oxygen species via superoxide dismutase and catalase revealed that redox state modulates enterobactin-driven changes in movement and lifespan through the secretin-family GPCR neuropeptide receptor PDFR-1. Together, these findings establish microbial enterobactin as a non-canonical neurotransmitter regulating neuronal mitochondrial function, behavior and aging, defining a gut bacteria-brain axis with broad implications for maintaining late-life health.

Metabolites are increasingly recognized as circulating molecules that regulate physiology, yet the mechanisms that couple intracellular production to organism-wide action remain poorly defined. Using the anorexigenic metabolite Lac-Phe as a tractable system, we identify the orphan transporter MCT6 (SLC16A5) as a physiologic intestinal Lac-Phe exporter. This mechanism controls the extent to which intracellularly synthesized Lac-Phe acquires systemic activity. MCT6 transports Lac-Phe, mediates its cellular efflux, and is required for maintaining its blood levels in mice following strong glycolytic stimuli. Both global and intestinal epithelial-specific deletion of MCT6 confers resistance to metformin-associated weight loss on a high-fat diet. Bypassing the transport defect with exogenous Lac-Phe normalizes the body weight phenotype of MCT6-KO mice. Together, these data connect MCT6 to metformin pharmacology and intestinal lactate metabolism, and more generally underscore the importance of transporter-mediated release in the conversion of an intracellular metabolic state into a circulating metabolite effector.

Mitochondrial remodelling is a key feature of neurogenesis in cortical and hippocampal neuronal lineages, yet the temporal control of organellar homeostasis in developing human motor neurons remains poorly understood. Here, we generated a high-resolution temporal resource integrating proteomics, metabolomics, and mitochondrial characterization across human iPSC-to-motor neuron differentiation. This analysis revealed prolonged mitochondrial depletion during early neurogenesis, characterized by reduced mitochondrial content, mitochondrial fragmentation, and lysosome-dependent turnover. This state persisted throughout the neural progenitor stage and was followed by recovery of mitochondrial content during neuronal maturation. During this transition, pyruvate utilization shifted from pyruvate carboxylation towards pyruvate dehydrogenase-dependent oxidation. Pharmacological manipulation of pyruvate partitioning demonstrated that this metabolic switch is required for human neuronal differentiation. Our findings establish a temporal framework for the metabolic control of human motor neuron differentiation and reveal a progenitor-specific state linking mitochondrial depletion to pyruvate metabolism.

Protein is unique among macronutrients due to its nitrogen content and that its building blocks, essential amino acids, cannot be synthesized by animals. Additionally, animals lack specialized amino acid storage sites, necessitating a continuous dietary intake. In the face of insufficient dietary protein, how do animals manage nitrogen limitation? Using Drosophila, we reveal a previously undocumented nitrogen flow from phospholipids to amino acids, a process enhanced by protein deprivation. Protein restriction triggers the degradation of phosphatidylethanolamine (PE), releasing the nitrogen-containing headgroup, ethanolamine, which subsequently serves a dual function. Ethanolamine stimulates protein intake by activating protein hunger neurons and, remarkably, along with phosphoethanolamine, donates nitrogen for amino acid biosynthesis. This nitrogen transfer is mediated by the microbiome and ethanolamine-phosphate phospho-lyase (ETNPPL) pathway in fly cells. Together, our findings identify phospholipids as a hidden nitrogen reservoir and reveal an unrecognized metabolic plasticity that reallocates nitrogen to maintain protein homeostasis.

Energy homeostasis and motivated behaviours are tightly interconnected processes coordinated by peripheral metabolic signals and central reward circuits. Among these signals, circulating triglycerides (TG) have emerged as neuromodulators of mesolimbic dopamine functions and reward processing. However, whether TG directly regulate the activity of defined neuronal populations within the nucleus accumbens (NAc) during appetitive behaviours remains unknown. Here, we investigated how central TG availability shapes the in vivo activity of NAc dopamine D2 receptor-expressing spiny projection neurons (D2R-SPNs) across distinct nutritional and metabolic states. Using in vivo fiber photometry in Drd2-Cre mice, we monitored D2R-SPN Ca2+ dynamics during appetitive Pavlovian conditioning and refeeding. D2R-SPNs progressively developed robust responses to a reward-predictive conditioned stimulus (CS+) over the course of learning, while their activity during reward consumption remained stable. Acute elevation of central TG levels through carotid infusion did not alter D2R-SPN activity or behavioural performance in lean chow-fed mice, irrespective of whether they were fed or food-restricted. In contrast, in mice exposed to a high-fat diet (HFD), central TG delivery suppressed cue-evoked D2R-SPN activity in food-restricted animals without affecting reward consumption or behavioural outputs. Likewise, TG significantly reduced refeeding-evoked D2R-SPN activation in fasted HFD-mice, while leaving novelty-induced neuronal responses unchanged. Together, these findings demonstrate that obesogenic conditions reveal a metabolic state-dependent sensitivity of accumbal D2R-SPNs to circulating lipids. This work identifies TG as context-dependent modulators of D2R-SPN function and uncovers a mechanism through which dietary history interacts with current metabolic state to reshape the neural encoding of reward-predictive cues and food-directed behaviours.

Neurons maintain specialized membranes necessary for lifelong circuit function. While neuronal membranes thus contain unique lipidomes, how and why specific lipids become synaptically enriched remains mysterious. Here, we identify a conserved hallmark of mature neurons, phosphatidylserines containing omega-3 fatty acids (n-3 PS), which dynamically accumulate during synaptogenesis from Drosophila to humans. This neuronal lipotype depends on dietary n-3 and lysosomal catabolism mediated by Saposin, the phospholipase PLA2G15, and the lysophospholipid transporter spin/SPNS1. Reducing neuronal PS biosynthesis by targeting the PS synthase (Pss) caused lipidomic, synaptic, and behavioral deficits exacerbated by impaired glycerophospholipid catabolism. While dietary, lysosomal, and lipoprotein manipulations broadly altered n-3 levels, silencing developmental activity-templating neurons selectively reduced n-3 PS. Remarkably, pathogenic Pss variants lacking feedback inhibition similarly reduced n-3 PS and impaired circuit function. As n-3 PS is reduced during neurodegeneration but absent from iPSC-derived neurons, important relationships between lipotype and physiology may not be recapitulated in vitro.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a common, clinically heterogeneous condition associated with liver-related and cardiometabolic morbidity. Sex differences in MASLD prevalence and progression are well recognized, but how they extend across the liver and visceral adipose tissue (VAT), a major source of lipid and endocrine signals to the liver, remains unclear. Here, we profiled paired liver and VAT biopsies from women and men with and without MASLD, generating single-nucleus datasets of 34 liver samples and 40 VAT samples, including 32 patient-matched pairs. In liver, women with MASLD showed a higher Kupffer-cell triglyceride-catabolism score (1.30-SD female-male difference; FDR=0.0022), accompanied by directionally consistent hepatocyte and endothelial programs. In VAT, women with MASLD showed higher adipocyte lipid and oxidative metabolism (0.87-1.15 SD; FDR<0.05 across seven pathways) together with macrophage immune activation, whereas men with MASLD showed stronger stromal communication routes. Across matched patients, gene modules linked VAT adipocyte and macrophage states to liver hepatocyte stress and lipid-receipt states. Bulk RNA-seq module scoring and paired liver-VAT analyses in 293 patients supported hepatic stress in both sexes, lower adipocyte signaling in women with increasing steatosis, and cross-tissue adipocyte-hepatocyte coupling (r=0.30; FDR=2.1e-6). Together, these findings identify distinct sex-associated transcriptional states in liver and VAT in MASLD and show that these states are coordinated across both tissues within the same individuals.

The hypothalamus is a central regulator of glucose and energy homeostasis, with arcuate nucleus (ARC) neurons orchestrating these processes. Agouti-related peptide (AgRP) and pro-opiomelanocortin (POMC) neurons integrate metabolic cues to control feeding behavior and systemic metabolism. Among these cues, fatty acids (FA) have emerged as key modulators of ARC neuronal activity. While neuronal sensing of circulating FA has begun to be defined, the contribution of FA derived from endogenous lipid stores to ARC neuron function and energy homeostasis remains largely unexplored. We recently identified lipid droplets (LD) as regulated FA reservoirs that control FA availability and metabolism in orexigenic AgRP neurons, thereby modulating their activity and regulating feeding. This prompted us to investigate whether LD-derived FA similarly regulate POMC neuron function. To this end, we targeted adipose triglyceride lipase (ATGL), which catalyzes the first committed step of LD lipolysis, in POMC neurons. We show that LD are present in POMC neurons under basal conditions both in vitro and in vivo, and that pharmacological or genetic inhibition of ATGL increases LD abundance. ATGL deficiency enhances spontaneous firing of POMC neurons and leads to reduced body weight, fat and lean mass in males, but not females. Consistent with enhanced glucose metabolism, ATGL loss lowers glycemia and insulinemia and increases carbohydrate utilization in chow-fed males. In contrast, ATGL deficiency does not alter metabolic adaptations to cold exposure, fasting or diet-induced obesity in either sex. Collectively, these findings establish ATGL-dependent LD lipolysis in POMC neurons as a previously unrecognized, sex-dependent regulator of energy homeostasis.

Cancer cachexia causes pathological weight loss, anorexia, and depressive symptoms, culminating in increased mortality. Using mouse models of pancreatic cancer cachexia, we identify tumor and hepatic inflammatory signaling, multi-organ energy charge depletion, and altered tryptophan metabolism. Metabolomic profiling and stable isotope tracing revealed altered tryptophan fates: serotonin depletion and kynurenine accumulation across tumor, liver, muscle and spleen. Kynurenine accumulation involved accelerated conversion of tryptophan to kynurenine with slowed downstream conversion to kynurenic acid. Restoring serotonin activity pharmacologically or metabolically normalized feeding, locomotion, and depression-like behavior, but wasting, tumor burden, and survival remained unchanged. Blocking kynurenine production or kynurenine-mediated AhR activation pharmacologically produced similar results. Thus, in cancer cachexia, the tryptophan pathway is subverted to deplete serotonin and accumulate kynurenine. This subversion contributes to cachexia-associated behavioral and feeding defects but not lethal systemic catabolism.

Neuropeptides mediate inter-tissue communication in energy metabolism, yet most remain functionally uncharacterized. Using C. elegans, we uncover a neuropeptide signaling cascade that drives lipid catabolism under chronic dietary restriction. We show that impaired pharyngeal function leads to dyspepsia and nutrient insufficiency, which activates a signaling axis comprising two peptide-receptor pairs, NLP-10-NPR-35 and FLP-18-NPR-4. This cascade originates in ADL sensory neurons, relays through multiple interneurons, and enhances triglyceride hydrolysis in the intestine. Genetic disruption of any component of this axis restores fat accumulation in diet-restricted mutants, whereas overexpression of individual components in well-fed animals is insufficient to drive fat mobilization. Importantly, the same neuropeptide axis is engaged in starved wild-type animals, indicating a general response toward energy deficit rather than mutant-specific effects. These findings reveal a conserved neuroendocrine circuit that senses internal energy shortage and orchestrates systemic metabolic adaptation, providing a conceptual framework for how neuropeptide networks integrate nutritional status with energy balance in animals.

Obesity-associated insulin resistance is driven in part by adipose tissue dysfunction and oxidative stress, yet the role of extracellular mechanisms in modulating insulin signaling remains poorly understood. Here, we show that extracellular Glutathione Peroxidase 3 (GPX3) enhances adipocyte insulin sensitivity and functionalities independent of its canonical peroxidase function. GPX3, a secreted selenoprotein highly expressed in white adipose tissue (WAT), is suppressed in obesity, but whether it actively regulates insulin sensitivity had not been established. We find that WAT GPX3 expression closely mirrors insulin receptor (InsR) expression across a broad range of metabolic parameters in humans and increases dynamically following weight loss. Prolonged therapeutic selenium supplementation in diet-induced obese mice selectively suppressed epididymal WAT oxidative damage, restored GPX3 and InsR expression, and enhanced insulin-stimulated AKT phosphorylation, improving oral glucose tolerance. Mechanistically, recombinant GPX3 extracellularly administered at physiological concentrations induced InsR expression across multiple adipocyte models, including pre-adipocytes, epididymal WAT explants, and adipocyte organoids, by promoting binding of the transcription factor Specificity Protein 1 (SP1) to the Insr promoter, while knockdown of GPX3 completely blocks SVF adipogenesis. These effects rely on the structural integrity, rather than selenocysteine-dependent activity. Thus, we identify a GPX3-SP1-InsR signaling axis in adipose tissue insulin sensitivity, suggesting GPX3 is a novel paracrine mediator of metabolic health and a potential therapeutic target to reinstate WAT insulin sensitivity in obesity and type 2 diabetes.

Folate is a driver of one-carbon metabolism that is essential in T cells. The cellular consequences of its deprivation in CD8+ T cells and impact on anti-tumor function are unknown. Single cell RNA-seq analysis of primary central nervous system lymphoma (PCNSL) patients' tumor samples revealed exhaustion in T cells with high folate metabolism gene expression signature. Functional genomics, metabolite profiling, and glycomics identified disrupted cell surface glycosylation as a major consequence of folate deprivation in CD8+ T cells. Mechanistically, folate deprivation reduced nucleotide sugar levels, perturbed glycosylation, and triggered the apoptotic unfolded protein response. Overexpressing folate transporters in adoptively transferred CD8+ T cells increased intratumoral T cell numbers and reduced exhaustion. Therefore, our data draw a mechanistic line between one-carbon metabolism, cell-surface glycosylation, and anti-tumor T cell function, and support a hypothesis that increased folate uptake can enhance CD8+ T cell function including ex vivo and in vivo CAR-T cells.

Parabrachial nucleus (PBN) CGRP neurons mediate aversive feeding suppression, but PBN circuitry governing homeostatic, non-aversive feeding suppression remains poorly understood. Here, we identify a circuit in which paraventricular hypothalamus (PVH) neurons innervate BDNF-expressing ventrolateral/dorsolateral PBN (vl/dlPBNBDNF) neurons, which project to multiple hypothalamic nuclei. Refeeding strengthens excitatory and weakens inhibitory synaptic transmission in LPBN-projecting PVH neurons and preferentially induced Bdnf and Fos in vl/dlPBNBDNF neurons compared with elPBNCGRP neurons, whereas noxious LPS preferentially activated elPBNCGRP neurons. Gain- and loss-of-function experiments demonstrated that vl/dlPBNBDNF neurons are sufficient and necessary for feeding suppression without inducing conditioned taste aversion and that this effect require BDNF-TrkB signaling. In addition to hypothalamic inputs, vl/dlPBNBDNF neurons receive dense inputs from major downstream targets of elPBNCGRP neurons. Thus, vl/dlPBNBDNF neurons define a non-aversive feeding-suppression circuit positioned to integrate hypothalamic homeostatic signals with aversive signals, thereby coordinating food intake with internal energy needs and potentially harmful physiological conditions.

Early embryogenesis relies extensively on maternally deposited products until zygotic genome activation, yet the dynamics for the synthesis of new proteins in mammalian embryos remains poorly characterized. To address this, we applied pulsed stable isotope labelling by amino acids in cell culture (pSILAC) combined with narrow-window data-independent acquisition mass spectrometry to single mouse oocytes and embryos to resolve de novo protein synthesis during early embryogenesis. This revealed that the maternal proteome is not a static reservoir, with components of the subcortical maternal complex and amino acid transporters SLC7A1/2 being actively synthesized during the earliest developmental stages. Furthermore, phosphoproteomic analysis identified hundreds of previously unreported phosphosites and extensive regulation during the oocyte-to-embryo transition. Notably, phosphorylation of the PRC2-interacting KLP motif of EZHIP emerged as a potential regulatory mechanism, with modification of this region reducing EZHIP-PRC2 interaction and coinciding with H3K27me3 remodelling. Together, single embryo pSILAC revealed a maternal proteome that is continuously synthesized, recycled, and post-translationally regulated during early embryogenesis.

Obesity is a major risk factor for interrelated cardiorenal metabolic diseases including type 2 diabetes, heart failure and chronic kidney disease (1-4). Although obesity develops when energy intake chronically exceeds energy expenditure, individuals vary markedly in the amount of adiposity and metabolic dysfunction that develop in response to comparable caloric excess. Controlled overfeeding studies demonstrate several-fold variation in weight gain between individuals (5), while susceptibility to metabolic dysfunction also differs substantially between populations (6), suggesting considerable variation in how excess nutrients are partitioned. Energy balance is conventionally considered in terms of intake and expenditure, yet energy-containing nutrients can also be eliminated from the body. The kidney is uniquely positioned to regulate this third component of energy balance by determining whether filtered metabolites are reabsorbed or excreted. The metabolic benefits of SGLT2 inhibition demonstrate that reducing renal nutrient reabsorption can favourably alter systemic metabolic function (7,8), but whether naturally occurring variation in renal nutrient conservation contributes to metabolic disease susceptibility remains unknown. Here, screening eleven genetically divergent mouse strains identified BXD34 mice as strikingly resistant to diet-induced obesity, ectopic lipid accumulation and insulin resistance despite increased caloric intake. Mechanistically, this protection was associated with widespread urinary nutrient loss accompanied by coordinated suppression of renal solute transport, but came at the cost of profound susceptibility to acute kidney injury. Together, these findings identify renal nutrient conservation as an underappreciated determinant of whole-body metabolism and reveal a renometabolic trade-off in which protection from obesity is achieved at the expense of renal resilience.

In animals, the transition from egg to embryo marks a rapid shift in cellular physiology, yet how mitochondria adapt to support this conversion remains poorly understood. We performed a targeted RNAi screen of conserved, nuclear-encoded mitochondrial proteins in Drosophila melanogaster to identify maternal effects. We identified distinct phenotypic classes affecting mitochondrial distribution or morphology in early embryos. We focused on Sideroflexin-1/3 (Sfxn1-3), a conserved mitochondrial inner membrane transporter, because it uniquely affects embryonic mitochondrial morphology. Maternal loss of Sfxn1-3 causes striking mitochondrial enlargement in embryos but not in oocytes, placing the trigger at the egg-to-embryo transition. Combining mitochondrial immunopurification with LC/MS-based metabolomic profiling, we show that amino acids are markedly enriched in Sfxn1-3-deficient mitochondria in both oocytes and embryos. This accumulation developmentally uncouples the metabolic and morphological phenotypes. By selectively disrupting discrete steps of egg activation, including translational activation, cell cycle re-entry, and fertilization, we demonstrate that mitochondrial swelling is independent of each, prompted instead by egg hydration. Beyond its reported role in proliferating cells as a serine importer, our data support a role for germline Sfxn1-3 in regulating inner mitochondrial membrane permeability to osmotically active amino acids. This activity is crucial for preserving organelle volume during the osmotic challenge of oocyte hydration. When this export fails, amino acid accumulation amplifies osmotic stress and drives rapid mitochondrial swelling. Together, these findings identify mitochondrial inner membrane permeability to amino acids as a key regulatory step that couples pre-established metabolite compartmentalization to osmotic adaptation at egg activation, and reveal how mitochondrial metabolic integrity and volume control are coordinated at this critical developmental transition.

The claustrum regulates reward processing, but its role in feeding remains unknown. We report that palatable food recruits claustral neurons and a claustrum-to-medial prefrontal cortex circuit whose activation reduces high-fat diet intake. These findings identify the claustrum as a key regulator of appetitive feeding, with potential implications for metabopsychiatric disorders.

Glucocorticoids (GCs) are regulators of metabolism, yet how physiological GC exposure remodels adipose metabolism and its relationship with circulating biomarkers remains poorly understood. In a randomised cross-over study of individuals with primary adrenal insufficiency, near-physiological GC exposure was compared to withdrawal using integrated serum metabolomics, abdominal subcutaneous microdialysis metabolomics, and adipose tissue transcriptomics. GC exposure induced coordinated changes in adipose lipid and amino-acid metabolism. Pathway-level entropy analysis revealed GC-state-dependent transcriptional reorganisation, with concentrated organisation during exposure and diffuse organisation during withdrawal. Integrative hypergraph and topological analyses revealed GC-state-dependent reorganisation of circulating metabolite-adipose tissue relationships, with greater higher-order coordination during exposure. The circulating phenylalanine/kynurenine ratio discriminated GC state, showed the strongest shared connectivity to GC-regulated adipose networks, persisted at midnight and in an independent dose-response study, and exhibited circadian rhythmicity. These findings reveal dynamic GC regulation of adipose metabolism and identify a circulating biomarker with potential to guide personalised GC therapy.

Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) arise, in part, from excessive free fatty acid flux from adipose tissue to the liver. Activin E (ActE, encoded by INHBE) suppresses adipocyte lipolysis through the type I receptor ALK7 (ACVR1C). Loss-of-function variants in INHBE and ACVR1C reduce waist-to-hip ratio in humans, yet genetic knockouts in mice produce insulin resistance and hepatic steatosis, suggesting discrepancies between human and mouse biology. We identify GDF3, a TGF-{beta} superfamily ligand upregulated in obese adipose tissue, as the principal endogenous antagonist of ActE/ALK7 signaling. Human transcriptomic datasets reveal coordinated dysregulation: hepatic INHBE expression and circulating ActE protein are elevated in obesity, while adipose ACVR1C is downregulated and GDF3 is reciprocally upregulated. Using ALK7-selective reporter assays, we show GDF3 inhibits ActE-driven SMAD2/3 signaling as a competitive antagonist rather than the weak agonist previously proposed. ActE suppressed beta-adrenergic-stimulated lipolysis in mouse and human adipocytes and primary human adipose tissue; GDF3 overexpression abolished this effect. In diet-induced obese mice, inducible Gdf3 deletion reduced adipose lipolysis, resolved hepatic steatosis and fibrosis, and improved insulin sensitivity, benefits abolished by Inhbe knockdown, confirming dependence on ActE signaling. Predicted loss-of-function variants in INHBE show only nominal, WHR-dependent associations with type 2 diabetes risk, potentially confounded by hematological effects on HbA1c. Gdf3 deficiency synergized with the clinical-stage anti-activin receptor antibody Bimagrumab to amplify fat-mass loss and improve glucose homeostasis in multiple MASH models. These findings establish GDF3 as an endogenous antagonist of ActE-ALK7 signaling and nominate GDF3 inhibition as a therapeutic strategy for MASLD/MASH.