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The role of metabolism in shaping enzyme structures over 400 million years.
No full textAdvances in deep learning and AlphaFold2 have enabled the large-scale prediction of protein structures across species, opening avenues for studying protein function and evolution1. Here we analyse 11,269 predicted and experimentally determined enzyme structures that catalyse 361 metabolic reactions across 225 pathways to investigate metabolic evolution over 400 million years in the Saccharomycotina subphylum2. By linking sequence divergence in structurally conserved regions to a variety of metabolic properties of the enzymes, we reveal that metabolism shapes structural evolution across multiple scales, from species-wide metabolic specialization to network organization and the molecular properties of the enzymes. Although positively selected residues are distributed across various structural elements, enzyme evolution is constrained by reaction mechanisms, interactions with metal ions and inhibitors, metabolic flux variability and biosynthetic cost. Our findings uncover hierarchical patterns of structural evolution, in which structural context dictates amino acid substitution rates, with surface residues evolving most rapidly and small-molecule-binding sites evolving under selective constraints without cost optimization. By integrating structural biology with evolutionary genomics, we establish a model in which enzyme evolution is intrinsically governed by catalytic function and shaped by metabolic niche, network architecture, cost and molecular interactions
The immunopathogenesis of autoimmune encephalitis-related psychosis, a comprehensive review of humoral and cellular mechanisms
No full textAutoimmune encephalitis has reshaped the understanding of neuropsychiatric disorders by highlighting the role of autoantibodies in psychosis symptoms, which often mimic primary psychosis conditions. This review synthesizes recent research on autoimmune encephalitis-related psychosis, broadening the focus from humoral immunity to T cell autoimmunity and the communication between the peripheral and central nervous systems. We discuss the identification of neuronal antigen targets, particularly the N-methyl-D-aspartate receptor (NMDAR), and their involvement in disease pathogenesis. Current treatments, such as plasma exchange and intravenous immunoglobulin, primarily target the pathogenicity of autoantibodies. However, emerging evidence suggests a crucial role for T cells, glia cell, and B cell in the immunopathogenesis of autoimmune encephalitis-related psychosis diseases. Autoimmune factors, including T and B cells, can either infiltrate the brain from the periphery or propagate via interacting with other cells, like glia, within the brain itself. This review advocates for a comprehensive approach to studying and treating these conditions, integrating both humoral and cellular mechanisms
Growth of the maternal intestine during reproduction.
No full textThe organs of many female animals are remodeled by reproduction. Using the mouse intestine, a striking and tractable model of organ resizing, we find that reproductive remodeling is anticipatory and distinct from diet- or microbiota-induced resizing. Reproductive remodeling involves partially irreversible elongation of the small intestine and fully reversible growth of its epithelial villi, associated with an expansion of isthmus progenitors and accelerated enterocyte migration. We identify induction of the SGLT3a transporter in a subset of enterocytes as an early reproductive hallmark. Electrophysiological and genetic interrogations indicate that SGLT3a does not sustain digestive functions or enterocyte health; rather, it detects protons and sodium to extrinsically support the expansion of adjacent Fgfbp1-positive isthmus progenitors, promoting villus growth. Our findings reveal unanticipated specificity to physiological organ remodeling. We suggest that organ- and state-specific growth programs could be leveraged to improve pregnancy outcomes or prevent maladaptive consequences of such growth.</p
Processed data and analysis scripts for the A-PREDICT trial
No full textThis repository contains processed data from Nanostring gene expression profiling and targeted panel DNA sequencing related to the A-PREDICT Clinical Trial.The analysis scripts included herein were utilized for the analyses presented in the manuscript titled "Blood Vessel Depletion and Immunomodulating Effects of Anti-VEGF Therapy in Metastatic Clear Cell Renal Cell Carcinoma: The Phase 2 A-PREDICT Trial." </p
The potent human CAR activator CITCO is a non-genotoxic hepatic tumour-promoting agent in humanised constitutive androstane receptor mice but not in wild-type animals.
No full textA large number of drugs and compounds produced by the chemical and agrochemical industry, often referred to as 'non-genotoxic carcinogens' (NGC), score as tumour promotors in rodent models. It is unclear whether these compounds act similarly in humans. The most extensively investigated compounds have been the anti-convulsive drugs, phenobarbital (PB), and phenytoin. Liver tumours induced by PB are dependent upon the activation of the constitutive androstane receptor (CAR). However, marked species differences in CAR activation by exogenous chemicals exist with some being much more potent activators of human CAR, e.g., 6-(4-chlorophenyl)imidazo[2,1-β][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl)oxime (CITCO). We have compared CITCO-induced tumour formation in the livers of mice in which murine CAR has been replaced with its human counterpart. Our findings reveal that CITCO-dependent liver tumours are only formed in mice-expressing human CAR and not in wild-type animals. In addition, contrary to one of the proposed mechanisms of NGC carcinogenicity, we show that CITCO did not induce a hyperplastic response in the liver of the humanised mice. These data raise some key questions about the mechanism of action of NGCs and identify the limitations of current rodent carcinogenicity test systems in relation to risk assessment.</p
DEK::NUP214 acts as an XPO1-dependent transcriptional activator of essential leukemia genes.
No full textThe t(6;9)(p22.3;q34.1) translocation/DEK::NUP214 fusion protein defines a distinct subgroup of younger AML patients classified as a separate disease entity by the World Health Organization. DEK is a nuclear factor with multifunctional roles, including gene regulation, while its fusion partner, NUP214, plays a pivotal role in nuclear export by interacting with transport receptors such as XPO1. However, the precise mechanism by which DEK::NUP214 drives leukemia remains unclear. A comprehensive multi-omics comparison of 57 AML primary samples (including whole genome sequencing, targeted sequencing, transcriptomics, and drug screening with >500 compounds) revealed that t(6;9) cases display a selective response to XPO1 inhibitors (Selinexor & Eltanexor) and a distinct transcriptomic signature characterized by the overexpression of FOXC1 and HOX genes that are key leukemia mediators. CUT&RUN experiments demonstrated the direct binding of DEK::NUP214 to the promoters of FOXC1 and HOXA/B clusters. Strikingly, the expression of these genes and the binding of DEK::NUP214 to their regulatory regions were selectively reduced upon XPO1 inhibition in t(6;9) cells. Altogether, these results identified a novel function of DEK::NUP214 as an XPO1-dependent transcriptional activator of key leukemia drivers and provide a rationale to explore the use of XPO1 inhibitors in this patient population
A multimodal imaging pipeline to decipher cell-specific metabolic functions and tissue microenvironment dynamics.
No full textTissue microenvironments are extremely complex and heterogeneous. It is challenging to study metabolic interaction between the different cell types in a tissue with the techniques that are currently available. Here we describe a multimodal imaging pipeline that allows cell type identification and nanoscale tracing of stable isotope-labeled compounds. This pipeline extends upon the principles of correlative light, electron and ion microscopy, by combining confocal microscopy reporter or probe-based fluorescence, electron microscopy, stable isotope labeling and nanoscale secondary ion mass spectrometry. We apply this method to murine models of hepatocellular and mammary gland carcinomas to study uptake of glucose derived carbon (13C) and glutamine derived nitrogen (15N) by tumor-associated immune cells. In vivo labeling with fluorescent-tagged antibodies (B220, CD3, CD8a, CD68) in tandem with confocal microscopy allows for the identification of specific cell types (B cells, T cells and macrophages) in the tumor microenvironment. Subsequent image correlation with electron microscopy offers the contrast and resolution to image membranes and organelles. Nanoscale secondary ion mass spectrometry tracks the enrichment of stable isotopes within these intracellular compartments. The whole protocol described here would take ~6 weeks to perform from start to finish. Our pipeline caters to a broad spectrum of applications as it can easily be adapted to trace the uptake and utilization of any stable isotope-labeled nutrient, drug or a probe by defined cellular populations in any tissue in situ.</p
Nitric oxide promotes cysteine N-degron proteolysis through control of oxygen availability.
No full textSelected proteins containing an N-terminal cysteine (Nt-Cys) are subjected to rapid, O2-dependent proteolysis via the Cys/Arg-branch of the N-degron pathway. Cysteine dioxygenation is catalyzed in mammalian cells by 2-aminoethanethiol dioxygenase (ADO), an enzyme that manifests extreme O2 sensitivity. The canonical substrates of this pathway in mammalia are the regulators of G-protein signaling 4, 5, and 16, as well as interleukin-32. In addition to operating as an O2-sensing mechanism, this pathway has previously been described as a sensor of nitric oxide (NO), with robust effects on substrate stability upon modulation of NO bioavailability being widely demonstrated. Despite this, no mechanism to describe the action of NO on the Cys/Arg N-degron pathway has yet been substantiated. We demonstrate that NO can regulate the stability of Cys N-degron substrates indirectly via the regulation of ADO cosubstrate availability. Through competitive, O2-dependent inhibition of cytochrome C oxidase, NO can substantially modify cellular O2 consumption rate and, in doing so, alter the availability of O2 for Nt-Cys dioxygenation. We show that this increase in O2 availability in response to NO exposure is sufficient to alter both dynamic and steady-state ADO substrate levels. It is likely that this mechanism operates to couple O2 supply and mitochondrial respiration with responses to G-protein-coupled receptor stimulation
Disruption of NANOG-driven epithelial-mesenchymal transition (EMT) and self-renewal restores drug sensitivity in colorectal cancer.
No full textAIMS: To investigate the regulatory role of NANOG in genes associated with stemness, symmetric division, and therapeutic resistance in colorectal cancer stem-like cells (CRC-SCs), with a focus on ERK/GSK-3β/β-catenin signalling and epithelial-mesenchymal transition (EMT), in order to evaluate the translational potential of targeting NANOG-associated signalling pathways. METHODS: Stemness, signalling activity, and cell division modes were analysed using 3D colonospheres enriched for CRC-SCs. Drug responses to the MEK inhibitor U0126 and the GSK-3β inhibitor TDZD-8 were assessed in CRC patient-derived organoids (PDOs), alongside molecular assays, immunohistochemistry with H-score quantification in xenograft models, and molecular dynamics simulations. RESULTS: NANOG overexpression enhanced the expression of stemness-associated genes, promoted symmetric cell division, and activated ERK/GSK-3β signalling, contributing to increased sphere formation. Inhibition of MEK and GSK-3β reduced EMT, cell proliferation, and symmetric division in CRC-SCs. NANOG-mediated dysregulation of ERK/GSK-3β altered β-catenin signalling and disrupted E-cadherin-dependent cell-cell adhesion. Molecular simulations and drug assays demonstrated that TDZD-8 and U0126 interfere with NANOG-DNA binding and β-catenin/E-cadherin interactions. CONCLUSIONS: NANOG drives CRC-SC maintenance via ERK/GSK-3β/β-catenin signalling and EMT modulation. This study offers significant insights into the translational impact of targeting NANOG and its downstream pathways with small-molecule inhibitors U0126 and TDZD-8 and presents a promising strategy to reduce CRC-SCs stemness, functionality, and tumourigenicity.</p
The interplay between innate immunity and nonimmune cells in lung damage, inflammation, and repair.
No full textAs the site of gas exchange, the lung is critical for organismal survival. It is also subject to continual environmental insults inflicted by pathogens, particles, and toxins. Sometimes, these insults result in structural damage and the initiation of an innate immune response. Operating in parallel, the immune response aims to eliminate the threat, while the repair process ensures continual physiological function of the lung. The inflammatory response and repair processes are thus inextricably linked in time and space but are often studied in isolation. Here, we review the interplay of innate immune cells and nonimmune cells during lung insult and repair. We highlight how cellular cross talk can fine-tune the circuitry of the immune response, how innate immune cells can facilitate or antagonize proper organ repair, and the prolonged changes to lung immunity and physiology that can result from acute immune responses and repair processes