Exposure to low levels of environmental challenges, known as hormetic stress, fosters subsequent stress resistance and promotes healthy aging in later life. However, specific mechanisms governing transcriptional reprogra...Exposure to low levels of environmental challenges, known as hormetic stress, fosters subsequent stress resistance and promotes healthy aging in later life. However, specific mechanisms governing transcriptional reprogramming upon hormetic nutrient stress remain elusive. Here, we identify histone H3 lysine 27 acetylation (H3K27ac) as a crucial driver of transcriptomic adaptation to hormetic fasting. Beyond its immediate function of enhancing lipid catabolism for alternative energy sources, stress-induced H3K27ac activates lifelong antioxidant defenses, thereby reducing reactive oxygen species (ROS) produced by stress-induced fatty acid oxidation and their accumulation during aging. Induced H3K27ac at metabolic genes, mediated by the pioneer factor PHA-4/FOXA, the cooperating transcription factor NHR-49/HNF4, and the nucleoporin 50 (NPP-16/NUP50), is crucial for lifespan extension under hormetic nutrient stress in Caenorhabditis elegans. Our findings establish H3K27ac as a key transcriptional switch bridging nutrient status with transcriptomic reprogramming, underpinning the longevity of hormetic fasting through orchestrating lipid catabolism and antioxidant defenses.
Selective removal of endoplasmic reticulum (ER) is important for cell health. Macroautophagy is the primary mechanism for the removal of the ER, but the ER can be cleared in a macroautophagy-independent manner. However,...Selective removal of endoplasmic reticulum (ER) is important for cell health. Macroautophagy is the primary mechanism for the removal of the ER, but the ER can be cleared in a macroautophagy-independent manner. However, the physiological relevance and mechanisms underlying macroautophagy-independent ER clearance remain largely unknown. Here we show that ER is cleared by lysosomes in a macroautophagy Atg gene-independent manner during development. This developmentally programmed Atg-independent ER clearance by lysosomes requires the ER protein Vap33 that promotes ER and lysosome contact. Oxysterol-binding protein (Osbp) is known to associate with Vap33, and Osbp lysosomal localization is required for ER clearance in cells lacking macroautophagy. Significantly, the cholesterol transport-associated protein Start1 regulates ER and lysosome contact, macroautophagy-independent ER clearance, and cholesterol transport from ER to the lysosome. These studies reveal that Vap33, Osbp, and Start1 promote ER clearance by lysosomes that is associated with cholesterol trafficking.
Hepatitis E virus (HEV) is the leading cause of viral hepatitis worldwide. We find that the HEV ORF2 capsid protein contains intrinsically disordered regions (IDRs) at its N terminus that promote condensate formation thr...Hepatitis E virus (HEV) is the leading cause of viral hepatitis worldwide. We find that the HEV ORF2 capsid protein contains intrinsically disordered regions (IDRs) at its N terminus that promote condensate formation through liquid-liquid phase separation both in vitro and in cellulo. Using correlative light and electron microscopy/tomography of HEV-infected cells, we observe that ORF2 forms higher order assemblies composed of orderly stacked tubular filaments resembling amyloid fibrils. By light microscopy, we detect ORF2 fibrils in multiple HEV-infected cell types, including human induced pluripotent stem cell-derived hepatocytes and neurons. Moreover, we discover that the central adapter protein TANK-binding kinase 1 is sequestered by ORF2 fibrils, thereby enabling immune evasion and enhancing viral replication. Together, our findings uncover a previously unrecognized mechanism by which HEV modulates host defense pathways and place HEV among the growing repertoire of viruses that exploit dynamic phase behavior as a broader strategy to regulate virus-host interactions.
Pancreatic cancer (PC) is a highly aggressive malignancy, with cancer stem cells (CSCs) playing a critical role in metastasis, therapy resistance, and recurrence, thereby presenting significant treatment challenges. Emer...Pancreatic cancer (PC) is a highly aggressive malignancy, with cancer stem cells (CSCs) playing a critical role in metastasis, therapy resistance, and recurrence, thereby presenting significant treatment challenges. Emerging evidence suggests that normal embryonic and adult progenitor stem cells share common gene signatures with CSCs. However, the early pluripotency-associated factors and stemness programs that become aberrantly reactivated during oncogenic transformation remain poorly defined. Here, we identify SLC22A3/OCT3 (Solute Carrier Family 22 Member 3) as a reactivated embryonic-associated signature that regulates PC stemness. SLC22A3 is markedly upregulated in CSC-enriched populations, and its silencing in PC cells grown in both 2D cultures and 3D organoid models significantly reduces key stemness features. Moreover, serotonin (5-HT) transport, mediated by SLC22A3, enhances overall stemness via downstream histone modifications, linking serotonin signaling to CSC regulation. Collectively, these findings establish SLC22A3 as a regulator of serotonin-driven stemness and a potential therapeutic target in PC.
Nocente MC, Della Rosa M, Malcolm AA
… +13 more, Lister G, Savin I, Ray-Jones H, Elderkin S, Tian R, Andrews S, Bendall A, Semprich CI, Kampmann M, Malysheva V, Rostovskaya M, Rugg-Gunn PJ, Spivakov M
Poised enhancers (PEs), co-marked by H3K4me1 and Polycomb-associated H3K27me3, are common in primed human pluripotent stem cells (hPSCs) resembling post-implantation epiblast but scarce in naive hPSCs modeling pre-implan...Poised enhancers (PEs), co-marked by H3K4me1 and Polycomb-associated H3K27me3, are common in primed human pluripotent stem cells (hPSCs) resembling post-implantation epiblast but scarce in naive hPSCs modeling pre-implantation epiblast. PEs form abundant chromosomal contacts with developmental genes, but the timing of their emergence, their relationship to enhancer poising, and their functional significance remain unclear. We devised high-resolution, PE-targeted Capture Hi-C to map PE contacts during the transition from naive to primed pluripotency. We find that enhancer poising emerges early in the transition, while the contacts show diverse dynamics. PROTAC-induced degradation of Polycomb repressive complex 2 early in the transition, but not inhibition of its H3K27 methyltransferase activity, weakens PE connectivity. Finally, PE contacts persist after developmental activation or ectopic CRISPRa targeting and can mediate long-range gene induction. Together, these findings reveal the temporal and mechanistic principles of PE connectivity and highlight a potential role of PE contacts in establishing developmental gene expression patterns.
Advanced amyloid beta (Aβ) pathology is associated with aberrant neuronal network activity and cognitive impairment in preclinical Alzheimer's disease (AD) models. Here, we assess Aβ pathology's impact on spatial informa...Advanced amyloid beta (Aβ) pathology is associated with aberrant neuronal network activity and cognitive impairment in preclinical Alzheimer's disease (AD) models. Here, we assess Aβ pathology's impact on spatial information processing in the medial entorhinal cortex (MEC) of 18-month App knock-in (APP KI) mice during exploration of open field arenas. Spatial information scores are decreased in APP KI MEC neurons versus age-matched controls. Border cell firing preferences are unstable across sessions and grid cell spatial periodicity is disrupted. Ratemap stability analysis using the Earth Mover's Distance indicates increased instability in spatially tuned APP KI neurons. Spatial decoding analysis indicates deficits in position and speed coding in APP KI mice across all comparisons. Additionally, APP KI mice display a mild hyperactive phenotype driven by narrow-spiking putative interneurons. These findings tie Aβ-associated dysregulation in neuronal firing to disruptions in spatial information processing that may underlie cognitive deficits associated with AD.
Protein-protein interactions underlie core brain functions, including neurotransmitter release, receptor activation, and intracellular signaling. Here, we present a compendium of protein-protein interactions conserved ac...Protein-protein interactions underlie core brain functions, including neurotransmitter release, receptor activation, and intracellular signaling. Here, we present a compendium of protein-protein interactions conserved across vertebrate brains, spanning over 300 million years of evolution. From 2,197 biochemical fractions across five vertebrate species, we identify over 81,000 high-confidence interactions among 6,108 conserved proteins. This interaction map (VerteBrain) reveals both regulatory and structural complexes, including extensive synaptonemal protein associations likely involved in inter-neuronal coordination. Conservation across species underscores essential roles in neuronal and glial function, as well as in additional tissues for more widely expressed complexes. The VerteBrain dataset uncovers candidate disease mechanisms, including roles for ARHGEF1 in short stature syndromes, synaptic vesicle trafficking complexes in epilepsy, and RELCH (Rab11-binding and LisH domain, coiled-coil and HEAT repeat-containing) in congenital deafness. VerteBrain provides a resource for investigating brain protein interactions and their relevance to human neurological disorders.
Immune cells elicit a continuum of transcriptional states after spinal cord injury (SCI). In mammals, inefficient debris clearance and chronic inflammation impede recovery and overshadow pro-regenerative immune functions...Immune cells elicit a continuum of transcriptional states after spinal cord injury (SCI). In mammals, inefficient debris clearance and chronic inflammation impede recovery and overshadow pro-regenerative immune functions. We found that zebrafish SCI elicits transient immune activation and efficient debris clearance. Transcriptomics and genetic ablation showed zebrafish macrophages are highly phagocytic and required for regeneration. Comparisons between zebrafish and mammalian macrophages identified transcription and immune response regulator (tcim) as an immune-enriched regenerative gene. Deletion of zebrafish tcim impairs phagocytosis and regeneration and activates a pro-inflammatory signature in leukocytes. Tcim expression in zebrafish and mouse macrophages establishes its conserved roles by promoting lipid metabolism and the reprogramming of myeloid precursors into activated phagocytes. This study underscores a requirement for elevated phagocytic capacity to achieve innate spinal cord repair and identifies a gene with conserved function in mammalian cells.
Fibroblast Growth Factor Receptors (FGFRs) are important cellular signaling mediators and are implicated in numerous cancers, including osteosarcoma (OS). Recent research shows that FGFR1 participates in inorganic phosph...Fibroblast Growth Factor Receptors (FGFRs) are important cellular signaling mediators and are implicated in numerous cancers, including osteosarcoma (OS). Recent research shows that FGFR1 participates in inorganic phosphate (Pi) sensing in osteoblasts, but how Pi activates FGFR1 remains unresolved. Here, we identify this unconventional mechanism of FGFR1 activation. We demonstrate that a large subset of OS cell lines is dependent on FGFR1 for growth, with Pi acting as an activating stimulus. In vivo experiments reveal that blood Pi levels are associated with tumor growth, implicating Pi-induced FGFR1 activation in OS progression. Elevated Pi triggers FGFR1 activation via Pi transport across the plasma membrane into mitochondria, increasing HO production. The HO burst oxidizes critical cysteine and methionine residues in FGFR1, promoting receptor activation and downstream signaling. These results reveal a targetable oncogenic mechanism for FGFR1 activation and explain the physiological Pi sensing mechanism.
Dendritic cells (DCs) are professional antigen-presenting cells that exhibit significant heterogeneity in development and function. This study found that among 17 cytokines screened, only IL-33 could induce DCs to displa...Dendritic cells (DCs) are professional antigen-presenting cells that exhibit significant heterogeneity in development and function. This study found that among 17 cytokines screened, only IL-33 could induce DCs to display a Th9-type cytokine profile, including IL-9, IL-4, and IL-13, in vitro. The response of DCs to IL-33 was synergistically enhanced by TGF-β and IL-4, defining a distinct subset termed DC9. Transcriptomic analysis revealed that DC9 possesses a unique gene expression signature, particularly in cytokine/chemokine clusters, compared to conventional DCs. Mechanistically, DC9 polarization depends on the JAK-STAT6-IRF4 signaling pathway. Functionally, DC9 preferentially drives naive CD4 T cell differentiation toward Th9 cells in vitro. In an ovalbumin-induced allergic airway inflammatory mouse model, DC9 cells were detectable in lungs and adoptive transfer of DC9 exacerbated disease severity, and increased pulmonary Th9 cells. Collectively, we identify DC9 as a novel DC subset induced by IL-33/TGF-β/IL-4 through the JAK-STAT6-IRF4 axis, which promotes Th9 differentiation and aggravates allergic airway inflammation.
Cognitive operations often require flexible implementation of stimulus-response contingencies, depending on context. We developed an olfactory task in which mice learned to associate a test odor with a directional lick r...Cognitive operations often require flexible implementation of stimulus-response contingencies, depending on context. We developed an olfactory task in which mice learned to associate a test odor with a directional lick response, conditional on a preceding context odor drawn from a different odor set. Two-photon imaging shows that the anterior lateral motor cortex (ALM) contains distinct populations encoding context, test odors, and choice. Optogenetic silencing during the context and delay periods impairs performance, suggesting that ALM contributes to configuring the appropriate contingency. Although context odors that instruct the same mapping are represented by separate populations, their influence converges at the level of choice-selective neurons. A subpopulation of these neurons exhibit dual selectivity for context and choice, forming what we term "contingency neurons." These findings suggest that ALM supports flexible behavior not by abstracting over context cues but by dynamically reconfiguring local circuits to route sensory input to the appropriate motor output.
Piezo2 is a force-gated ion channel that functions as a sensor of mechanical touch, proprioception, lung inflation, and gut transit. Human Piezo2 contains seven domains that are alternatively spliced in a tissue-specific...Piezo2 is a force-gated ion channel that functions as a sensor of mechanical touch, proprioception, lung inflation, and gut transit. Human Piezo2 contains seven domains that are alternatively spliced in a tissue-specific fashion resulting in the expression of at least 22 distinct variants. Despite the relevance of Piezo2 in human physiology, its sensitivity to membrane tension, and how this fundamental biophysical property is affected by alternative splicing, are unknown. Here, we use cell-attached pressure-clamp electrophysiology combined with differential interference contrast microscopy to quantify the response of Piezo2 to membrane tension and identify the alternatively spliced exon 35 as a domain sufficient to confer high sensitivity to membrane tension and cellular indentation. We further show that physiological variants of Piezo2 sense mechanical forces with distinct sensitivities and dynamic ranges. Together, our findings rationalize how Piezo2 variants may fulfill distinct physiological functions required for somatosensation and interoception.
5-methylcytosine is a reversible regulatory mark in mammalian genomes whose distribution varies across genetic loci, cell types, and disease conditions. Cytosine-5 methylation is brought about by three AdoMet-dependent D...5-methylcytosine is a reversible regulatory mark in mammalian genomes whose distribution varies across genetic loci, cell types, and disease conditions. Cytosine-5 methylation is brought about by three AdoMet-dependent DNA methyltransferases (DNMTs) with primary tasks in de novo (DNMT3A and DNMT3B) or maintenance (DNMT1) methylation. However, the precise roles and interplay between the three methylation "writers" remain obscure. To selectively track an individual DNMT, we derived a heterozygous mESC line with chromosomal alleles expressing the wild-type and a sterically engineered variant of DNMT1. This enables catalysis-dependent chemical tagging DNMT1 methylation sites upon in-cell delivery of a synthetic AdoMet analog. Genome-wide mapping of the DNMT1 catalysome during naive-to-primed transition of mESC shows that DNMT1 contributes to de novo methylation of germline genes and competes with active demethylation at boundaries of CpG islands. Altogether, we provide a unique tool for chemical monitoring of individual methylation writers in minimally perturbed developmental and disease models.
Bai D, Sun J, Chen C
… +17 more, Jia Y, Li Y, Liu K, Zhang Y, Yin J, Liu Y, Han X, Ruan J, Kou X, Zhao Y, Wang H, Wang Z, Chen M, Teng X, Jiang C, Gao S, Liu W
Multiple sclerosis (MS) is a complex immune-mediated disorder with polygenic and multicellular underpinnings, necessitating cell-type-specific molecular studies to delineate dysregulated pathways. Here, we profile 1,075...Multiple sclerosis (MS) is a complex immune-mediated disorder with polygenic and multicellular underpinnings, necessitating cell-type-specific molecular studies to delineate dysregulated pathways. Here, we profile 1,075 transcriptomes from 167 patients with MS and 42 healthy participants across six peripheral immune cell-type-states. MS-associated transcriptional differences are more pronounced in primary (unstimulated) immune cells than in in vitro-stimulated counterparts. We identify shared and cell-type-specific transcriptional alterations at the level of genes, pathways, and co-expressed gene modules, prioritizing regulators, such as ZBTB16, across T cells and monocytes, and replicating six MS-associated modules in independent datasets. The top T cell module is enriched for MS susceptibility genes and affects proliferation. The top monocyte module implicates dysregulated TNF-α/NF-κB signaling, for which an in silico drug screen and in vitro validation nominate alvespimycin as a candidate modulator. Together, these findings define stable peripheral immune dysregulation signatures in MS that may serve as diagnostic or prognostic biomarkers in at-risk individuals.
Spatial regulation of mRNA polyadenylation emerges as a key mechanism shaping cellular function. The TENT5/FAM46 family comprises four non-canonical poly(A) polymerases that stabilize transcripts encoding ER-targeted pro...Spatial regulation of mRNA polyadenylation emerges as a key mechanism shaping cellular function. The TENT5/FAM46 family comprises four non-canonical poly(A) polymerases that stabilize transcripts encoding ER-targeted proteins, with mutations linked to diseases of professional secretory cells. Using transcriptomic and proteomic profiling with systematic mutagenesis, we show how paralog-specific divergence drives distinct localization, interactions, and functions. TENT5D, the most ER-associated member, remodels the proteome, enhancing the expression of ER, ERGIC, Golgi, and lysosomal proteins. In contrast, TENT5B lacks ER targeting and regulates proteins involved in cell division. A member-specific C-terminal region that binds ER-transmembrane FNDC3 proteins is necessary and sufficient for ER localization. Mutations in this region of TENT5C, found in multiple myeloma, impair FNDC3 binding or stability, reducing immunoglobulin production and tumor-suppressive activity. Overall, we define a domain-encoded mechanism linking TENT5 localization to transcript selectivity and secretory output, with direct implications for compartmentalized mRNA regulation and development of RNA-based therapeutic strategies.
During Mycobacterium tuberculosis (Mtb) infection, infected alveolar macrophages (AMs) initially up-regulate a nuclear factor erythroid 2-related factor 2 (NRF2)-regulated cell-protective program, which is detrimental to...During Mycobacterium tuberculosis (Mtb) infection, infected alveolar macrophages (AMs) initially up-regulate a nuclear factor erythroid 2-related factor 2 (NRF2)-regulated cell-protective program, which is detrimental to host control and impedes AM activation, including MHC II expression. MHC II is critical for CD4 T cell activation and host immunity during Mtb infection. We hypothesized that NRF2 regulates the MHC II pathway and AM antigen presentation to T cells. We found that NRF2 inhibits MHC II, but not MHC I, specifically in AMs, following Mtb infection in vitro and in vivo. NRF2 dampens Ciita and H2-Ab1 gene expression in uninfected AMs, and MHC II inhibition by NRF2 is retained following innate stimuli and IFNγ exposure. NRF2 expression in Mtb-infected AMs impedes their ability to activate ESAT6-specific CD4 T cells. Thus, although NRF2 expression enhances cell-protective functions, it has the unexpected consequence of limiting innate-adaptive crosstalk, which can impair CD4 T cell activation and host immunity during Mtb infection.
The medial prefrontal cortex (mPFC) weighs options during decision-making, but its role in inferring competing emotional states and how this process changes with age, remain unclear. We recorded brain activity with funct...The medial prefrontal cortex (mPFC) weighs options during decision-making, but its role in inferring competing emotional states and how this process changes with age, remain unclear. We recorded brain activity with functional MRI while 20 young and 40 older adults inferred the emotions of actors shown in ecological yet controlled social interactions. Socially shared representations about the timing, accuracy, and uncertainty of emotional inferences were defined using response distributions from an independent sample viewing the same scenes. mPFC activity is sensitive to emotional inference and scales with the social uncertainty of alternative emotional hypotheses. Social consensus regarding the emotional conflict elicited by the actors' interaction is associated with increased representational similarity in the mPFC, with this effect being modulated by age. Together, these results indicate that the mPFC integrates the variability of social representations to infer emotions in others and provide an account of age-related changes in social cognition.
Bracey NA, Beppler C, Bilich T
… +11 more, Long AH, Sola E, Barak A, Few-Cooper TJ, Mohsin A, Shankar V, Mallajosyula V, Kamalyan L, Capasso R, Shen-Orr SS, Davis MM
A decline in specific antibody responses is a hallmark of human aging, yet the differential contributions of B and T lymphocytes remain unclear. CXCL13 is a chemokine that shapes germinal center (GC) organization, but th...A decline in specific antibody responses is a hallmark of human aging, yet the differential contributions of B and T lymphocytes remain unclear. CXCL13 is a chemokine that shapes germinal center (GC) organization, but the regulation of human-specific CXCL13 T follicular helper (Tfh) cells during aging is not known. Using human tonsil organoids, single-cell RNA sequencing, and CRISPR perturbations, we mapped age-associated changes in Tfh cells, the cell type that provides help to B cells in GCs. Tonsil organoids from older donors generate weaker influenza-specific antibody responses, which we trace to Tfh cell defects rather than B cells. Single-cell profiling revealed a selective loss of mature CXCL13 GC-Tfh cells accompanied by accumulation of precursor states. Trajectory analysis shows that aging arrests Tfh maturation at the early activated precursor transition, and CRISPR perturbations identify BACH2 and SOX4 as regulators of differentiation reduced with age. These findings reveal a human-specific mechanism of immune aging with implications for strategies to restore humoral immunity.