ABSTRACT:Excess iron induces tissue toxicity in various conditions, including hereditary hemochromatosis (HH). Hepcidin, a liver-derived hormone encoded by the HAMP gene, plays a pivotal role in regulating systemic iron by mediating the degradation of ferroportin (FPN), the sole cellular iron exporter. Previous research found that the E3 ubiquitin ligase RNF217 is a key regulator of iron homeostasis by directly affecting FPN degradation; however, the role of RNF217 overexpression in iron-overload disorders such as HH is poorly understood. To address this question, we generated both global and intestine-specific Rnf217-overexpressing mice and then crossed these mice with hemojuvelin knockout (Hjv-/-) mice, a model for studying hemochromatosis. We found that both global and intestine-specific Rnf217 overexpression caused an identical rescue of the HH phenotype, implicating duodenal enterocytes as the main site where RNF217 overexpression exerts its beneficial effects. Moreover, we found that intestinal Rnf217 overexpression significantly reduced iron accumulation in the serum and in vital organs; importantly, these effects were not correlated with hepcidin levels. In summary, our findings demonstrate that intestinal RNF217 overexpression can directly suppress iron absorption by modulating FPN protein levels, bypassing hepcidin. This suggests a possible therapeutic strategy for iron-overload disorders, warranting further study to establish its clinical potential.

Fine-tuning of energy metabolism is essential for the survival and suppressive function of regulatory T cells (Treg cells). Here, we show that Treg cells with a high energetic state display enhanced functional capacity. Using a screen of mitochondrial inhibitors, we identified copper chelators and ionophores as modulators of Treg cell energetic state. T cell receptor (TCR) stimulation in vitro and human autoimmune conditions increased the labile copper pool in Treg cells. In murine Treg cells, we characterized Slc31a1 as a major copper transporter that supports oxidative phosphorylation, sustains nicotinamide adenine dinucleotide/reduced NAD+ (NAD+/NADH) homeostasis, and promotes histone acetylation at loci encoding core Treg cell functional molecules. These mechanisms collectively ensured energy production and Treg cell functionality, which were indispensable for peripheral immune tolerance but could be rescued by the copper ionophore elesclomol. Together, our findings identify copper metabolism as a critical regulator of Treg cell functionality and suggest potential therapeutic avenues for autoimmune diseases.

Niu et al.1 introduce SPTEdU-seq, a strategy that integrates spatial transcriptomics with EdU-based temporal labeling to jointly capture transcriptional states and proliferative history. This approach provides a framework for resolving spatiotemporal cell dynamics in development, regeneration, and cancer.

Lewy bodies, a pathological hallmark of Parkinson's disease, are α-synuclein-enriched cytoplasmic inclusions that drive progressive neurodegeneration. A long-standing yet unmet goal has been the visualization of α-synuclein (α-Syn) inclusions in live brain and measurements of their pathological effects on individual neurons. Here, we developed genetically encoded reporters and knock-in mouse lines to achieve this goal. The reporters exhibited a 5-fold increase in fluorescence upon incorporation into α-Syn inclusions. They reliably reflected α-Syn inclusion propagation in the cortex of awake mice. Coupled with Ca2+ imaging and whole-cell recording, the reporters enabled measurement of the pathological effects of inclusions on neuronal activity and synaptic function. They could be selectively targeted to specific neuronal subtypes, facilitating measurement of the pathological effects on transcriptomes and metabolomes at the single-cell level. In live-cell imaging, the reporters helped identify inhibitors of α-Syn inclusion formation. Collectively, these genetically encoded reporters support multiple applications to study α-Syn inclusions in live brain.

Ferroptosis is an iron-dependent form of regulated cell death driven by disrupted iron homeostasis and uncontrolled lipid peroxidation. Various metabolites and enzymes regulate cellular sensitivity to ferroptosis by affecting iron, lipid and redox metabolism. These pathways not only signal ferroptotic cell death but also affect the biology of T cells. The pathways include mechanisms by which iron metabolism regulates T cell activation via transferrin receptor 1-mTOR signalling, mechanisms by which lipid peroxidation drives vulnerability to ferroptosis in tumour-infiltrating CD8+ T cells, and mechanisms by which redox networks are balanced to maintain T cell survival. Here, we highlight the T cell subset-specific effects of ferroptosis-related pathways and ferroptosis susceptibility, and the implications for immunotherapy. We also discuss the emerging therapeutic strategies, including ferroptosis-resistant adoptive T cell therapy and ferroptosis-inducing approaches, that enhance the efficacy of immune checkpoint blockade for cancer treatment. Finally, we propose a framework for precision T cell-based immunotherapies, positioning ferroptosis as a tunable node linking T cell biology to clinical innovations.

Elucidating how microscale molecular architecture shapes macroscale brain network organization requires integrative frameworks that link transcriptional profiles, structural connectivity, and functional dynamics across scales. However, existing evidence largely relies on cross-modal data acquired from different individuals, limiting causal and mechanistic inference. To overcome this, we performed concurrent awake optogenetic functional MRI (opto-fMRI) and spatial transcriptomics (opto-ST) within the same mice, enabling coupled analysis of neural activity and gene expression in fronto-thalamic and hippo-thalamic networks. Optogenetic stimulation of the medial prefrontal cortex (mPFC) or subiculum (SUB) evoked distinct thalamic BOLD response patterns. Multivariate spatial modeling revealed that energy metabolism and core biochemical processes underlie these GLM-derived activation patterns-a finding further supported by data-driven PCA, which identified enrichments in similar biological processes. These results suggest that conventional linear models predominantly capture linear gene-process relationships, highlighting the need for nonlinear computational approaches in fMRI analysis. Overall, our multi-omics framework reveals that genes regulating metabolic pathways underlie region-specific neurovascular coupling, thereby bridging local transcriptomics and large-scale functional imaging signatures.

Copper dysregulation has been linked to human health, disorders, and hematopoiesis. However, the underlying mechanisms remain elusive. Here, we demonstrate the pivotal role of dietary copper via the transporter Slc31a1(Ctr1) in copper homeostasis, but not cuproptosis, during postnatal hematopoiesis. Specifically, Slc31a1-mediated copper uptake sustains the differentiation and commitment of multipotent progenitors from short-term hematopoietic stem cells (HSCs). Using transcriptomic analyses, we reveal a disrupted differentiation program in hematopoietic stem and progenitor cells (HSPCs) in diet-induced copper-deficient mice or hematopoietic-specific Slc31a1 knockout (vKO) mice. Further, we show that Slc31a1 and copper are indispensable for sustaining mitochondrial activity via regulating Mtco1 and Mtco2 (subunits of Complex IV) within HSPCs. Notably, we show that the chemical compound elesclomol, also well-known as a potent cuproptosis agonist, significantly alleviates severe anemia and partially recovers HSPC mitochondrial function in vKO mice via its activity as a copper ionophore, but with no effect on cuproptosis. We thus renamed elesclomol as CupriActivitor1(CuA1), which is a more specific and descriptive term. These findings demonstrate the critical role and mechanism of copper, Slc31a1, and CuA1 in maintaining HSC homeostasis via modulation of mitochondrial energy metabolism. The study sheds light on the molecular basis of HSC fate decisions by copper or CuA1 and opens new avenues for the development of novel therapeutic strategies for copper-related disorders and blood diseases. Given the critical and multifaceted nature of copper, we propose establishing a novel interdisciplinary field termed "Cuprology". This discipline will advance our understanding of copper's roles in physiological and pathological processes.

Asthma is a chronic inflammatory airway disease characterized by defective epithelial repair, resulting from metabolic dysregulation in facultative progenitor cells. Here, we investigate how pyruvate metabolism in airway club cells controls epithelial differentiation and allergic airway inflammation. Single-cell transcriptomics revealed elevated glycolytic activity in club and goblet cells from patients with asthma. In an ovalbumin (OVA)-induced asthma model, conditional deletion of Mpc2—but not Ldha—in club cells impaired club-to-goblet cell differentiation, reduced CLCA3 and Foxa3 expression, and attenuated eosinophilic inflammation and Il-13 expression. Mpc2 loss increased Cxcl17 expression in club cells, promoting Cxcl17-Cxcr4 signaling with alveolar macrophages that suppressed CCL17-mediated type 2 inflammation. Neutralizing CCL17 phenocopied the Mpc2 knockout by reducing airway inflammation and goblet cell differentiation. These findings reveal a metabolic-immune crosstalk underlying asthma pathogenesis and identify mitochondrial pyruvate metabolism as a therapeutic target to limit epithelial remodeling and type 2 inflammation.

Unraveling the cellular and molecular characteristics of human prefrontal cortex (PFC) development is crucial for understanding human cognitive abilities and vulnerability to neurological and neuropsychiatric disorders. Here, in this study, we created a comparative repository for gene expression, chromatin accessibility and spatial transcriptomics of human and macaque postnatal PFC development at single-cell resolution. Integrative analyses outlined species-specific dynamic trajectories of different cell types, highlighting key windows and gene regulatory networks for processes such as synaptogenesis, synaptic pruning and gliogenesis. We identified regulatory correlates of the prolonged development of human PFC relative to macaques. Glial progenitors showed higher proliferation capability in humans compared to macaques, associated with distinct gene expression profiles. Furthermore, we uncovered cell types and lineages most susceptible to neurodevelopmental and neuropsychiatric disorders, focusing on transcription factors with human-specific expression features. In summary, our discoveries shed light on human-specific regulatory programs extending postnatal cortical maturation through coordinated neuronal and glial development, with implications for cognition and neurodevelopmental disorders.

Floral scent is a crucial characteristic that significantly influences reproductive processes and indicates the ornamental value of many plants. Antique Lotus (Nelumbo), an important ornamental germplasm, has high archaeological and cultural value in China. Although many studies have examined this plant, the floral fragrance characteristics remain unexplored. This study analyzed floral volatile profiles in three floral organs from six Antique Lotus. A combination of dynamic headspace collections and gas chromatography-mass spectrometry analyses revealed a total of 64 volatile components, comprising 38 terpenoids, two benzenoids/phenylpropanoids, and 24 fatty acid derivatives. The stamens were found to contain the greatest number of volatile compounds, while petals exhibit the highest content. Fatty acid derivatives were the primary volatile substances in petals, while stamens and receptacles were dominated by benzenoids/phenylpropanoids. These findings not only lay a foundation for aroma breeding but also provide a theoretical basis for the resource development of Antique Lotus.

Multicellular organisms rely on inter-organ communication networks to maintain vital parameters within a dynamic physiological range. Macrophages are central to this homeostatic control system, sensing deviations of those parameters and responding accordingly to support tissue function and organismal homeostasis. Here we demonstrate that dysregulation of iron metabolism in parenchyma cells, imposed by the deletion of ferritin H chain, is sensed by monocyte-derived macrophages. In response, macrophages derived from circulating monocytes support tissue function, energy metabolism and thermoregulation, as demonstrated in bone marrow chimeric and parabiotic mice. This salutary effect is contingent on a transcriptional program, controlled in macrophages by the transcription factor A mitochondria. This transcriptional response acts in a non-cell autonomous manner to support the mitochondria of parenchyma cells, irrespectively of mitochondrial transfer. In conclusion, monocyte-derived macrophages cross-regulate Fe and energy metabolism to support tissue function and organismal homeostasis. ### Competing Interest Statement The authors have declared no competing interest.

CLDN4 belongs to a multi-transmembrane protein family of claudins, which mainly functions in cell-cell adhesion and migration. MicroRNAs (miRNAs) are important post-transcriptional regulating factors that participate in broad biological process including immunity. Through high-throughput RNA sequencing strategy, a flounder miRNA, miR-29-x, was identified to be responsible to both bacteria and virus. In this study, we explored the regulatory mechanism and function of miR-29-x and its target gene of flounder CLDN4 (named PoCLDN4). We proved that miR-29-x could interact with the 3'UTR of PoCLDN4 and negatively regulate its expression. PoCLDN4 located on cell membrane, while the depletion of extracellular loop E2 abolished the membrane localization of this protein. E3 could bind different bacteria, and mutation of the amino acids of 13E and 18E enhanced this capacity, while mutation of 10L abolish this capacity. Further study revealed the bacteria killing effect of E3 and verified 10L as a key factor. These results identified the interaction between miR-29-x and PoCLDN4, and unraveled the function as well as the molecular basis of flounder CLDN4 in anti-bacterial immunity.

Elucidating the regulatory mechanisms underlying the development of different brain regions in humans is essential for understanding advanced cognition and neuropsychiatric disorders. However, the spatiotemporal organization of three-dimensional (3D) chromatin structure and its regulatory functions across different brain regions remain poorly understood. Here, we generated an atlas of high-resolution 3D chromatin structure across six developing human brain regions, including the prefrontal cortex (PFC), primary visual cortex (V1), cerebellum (CB), subcortical corpus striatum (CS), thalamus (TL), and hippocampus (HP), spanning gestational weeks 11–26. We found that the spatial and temporal dynamics of 3D chromatin organization play a key role in regulating brain region development. We also identified H3K27ac-marked super-enhancers as key contributors to shaping brain region-specific 3D chromatin structures and gene expression patterns. Finally, we uncovered hundreds of neuropsychiatric GWAS SNP-linked genes, shedding light on critical molecules in various neuropsychiatric disorders. In summary, our findings provide important insights into the 3D chromatin regulatory mechanisms governing brain region-specific development and can serve as a valuable resource for advancing our understanding of neuropsychiatric disorders.

Despite advances in whole-brain imaging technologies, the lack of quantitative approaches to bridge rodent preclinical and human studies remains a critical challenge. Here we present TransBrain, a computational framework enabling bidirectional translation of brain-wide phenotypes between humans and mice. TransBrain improves human-mouse homology mapping accuracy through (1) a cortical and subcortical detached region-specific deep neural network trained on integrated multimodal human transcriptomics to improve cortical correspondence (89.5% improvement over the original transcriptome), which revealed 2 evolutionarily conserved gradients, and (2) a graph-based approach to construct a unified cross-species representational space incorporating anatomical hierarchies and structural connectivity. We demonstrate TransBrain's utility through three cross-species applications: quantitative assessment of resting-state brain organizational features, inferring human cognitive functions from mouse optogenetic circuits and translating molecular insights from mouse models to individual-level mechanisms in autism. TransBrain enables quantitative cross-species comparison and mechanistic investigation of both normal and pathological brain functions.

In clinics, hepcidin levels are elevated in various anemia-related conditions, particularly in iron-refractory anemia and in high inflammatory states that suppress iron absorption, which remains an urgent unmet medical need. To identify effective treatment options for various types of iron-refractory anemia, the potential effect of hypoxia and pharmacologically-mimetic drug FG-4592 (Roxadustat) are evaluated, a hypoxia-inducible factor (HIF)-prolyl hydroxylase (PHD) inhibitor, on mouse models of iron-refractory iron-deficiency anemia (IRIDA), anemia of inflammation and 5-fluorouracil-induced chemotherapy-related anemia. The potent protective effects of both hypoxia and FG-4592 on IRIDA as well as other 2 tested mouse cohorts are found. Mechanistically, it is demonstrated that hypoxia or FG-4592 could stabilize duodenal Hif2α, leading to the activation of Fpn transcription regardless of hepcidin levels, which in turn results in increased intestinal iron absorption and the amelioration of hepcidin-activated anemias. Moreover, duodenal Hif2α overexpression fully rescues phenotypes of Tmprss6 knockout mice, and Hif2α knockout in the gut significantly delays the recovery from 5-fluorouracil-induced anemia, which can not be rescued by FG-4592 treatment. Taken together, the findings of this study provide compelling evidence that targeting intestinal hypoxia-related pathways can serve as a potential therapeutic strategy for treating a broad spectrum of anemia, especially iron refractory anemia.

Cell segmentation is the first step in parsing spatial transcriptomic data, often a challenging task. Existing cell segmentation methods do not fully leverage spatial cues between nuclear images and transcripts, tending to produce undesirable cell profiles for densely packed cells. Here, we propose CellCUT to perform cell segmentation and transcript assignment without additional manual annotations. CellCUT provides a flexible computational framework that maintains high segmentation accuracy across diverse tissues and spatial transcriptomics protocols, showing superior capabilities compared to state-of-the-art methods. CellCUT is a robust model to deal with undesirable data such as low contrast intensity, localized absence of transcripts, and blurred images. CellCUT supports a human-in-the-loop workflow to enhance its generalizability to customized datasets. CellCUT identifies subcellular structures, enabling insights at both the single-cell and subcellular levels. ### Competing Interest Statement The authors have declared no competing interest.

Lissencephaly is a rare brain malformation for which our understanding remains limited due to the absence of suitable animal models that accurately represent human phenotypes. Here, we establish doublecortin (DCX) knockout ferrets as a model that faithfully replicates key features of the disorder. We reveal the critical roles of DCX in neural progenitor cell proliferation and radial glial fiber extension, processes essential for normal cortical development. Utilizing single-nucleus RNA sequencing (snRNA-seq) and spatial transcriptomics, we provide a detailed atlas of the lissencephalic cortex, illustrating disrupted neuronal lamination and the specific interactions between inhibitory and excitatory neurons. These findings enhance our understanding of the cellular and molecular mechanisms underlying lissencephaly and highlight the potential of DCX knockout ferrets as a valuable tool for neurodevelopmental research, offering insights into both the pathology of lissencephaly and the general principles of brain development.

Unlike megabats, which rely on well-developed vision, microbats use ultrasonic echolocation to navigate and locate prey. To study ultrasound perception, here we compared the auditory cortices of microbats and megabats by constructing reference genomes and single-nucleus atlases for four species. We found that parvalbumin (PV)+ neurons exhibited evident cross-species differences and could respond to ultrasound signals, whereas their silencing severely affected ultrasound perception in the mouse auditory cortex. Moreover, megabat PV+ neurons expressed low levels of complexins (CPLX1-CPLX4), which can facilitate neurotransmitter release, while microbat PV+ neurons highly expressed CPLX1, which improves neurotransmission efficiency. Further perturbation of Cplx1 in PV+ neurons impaired ultrasound perception in the mouse auditory cortex. In addition, CPLX1 functioned in other parts of the auditory pathway in microbats but not megabats and exhibited convergent evolution between echolocating microbats and whales. Altogether, we conclude that CPLX1 expression throughout the entire auditory pathway can enhance mammalian ultrasound neurotransmission.