Biology subjects

Kim, M.

Publications and source records attributed to Kim, M..

At least 109 records · Page 6Linked to original sources

bioRxiv2025

Systematic Growth Factor Profiling Platform for 3D Tumor Models Reveals Estradiol-Responsive Cellular Mechanisms of Immunotherapy Resistance

Current organoid culture systems face critical limitations: standardized growth factor formulations fail to capture patient-specific signaling requirements, and single-cell-type approaches overlook tumor-stromal interactions essential for understanding immunotherapy resistance. To address these challenges, we developed an innovative biofabrication platform that systematically integrates patient-derived three-dimensional (3D) cultures with comprehensive growth factor profiling across 128 combinations. Through systematic screening of 23 ovarian cancer patient samples and single-cell RNA sequencing, we identified two estradiol-responsive cellular populations that coordinate immunosuppression: a malignant cell fraction (MAL.PDCD5) and a cancer-associated fibroblast (CAF) fraction (FB.TNFSF10). MAL.PDCD5 cells suppress immune infiltration by downregulating antigen presentation, while FB.TNFSF10 cells promote immunosuppression through enhanced TGF-{beta} signaling. Spatial transcriptomic analysis revealed striking mutual exclusivity between FB.TNFSF10 cells and T/NK cells, providing evidence of active immune cell exclusion. Most significantly, FB.TNFSF10 abundance emerged as a robust predictor of immune checkpoint inhibitor therapy resistance across multiple cancer cohorts, independent of conventional biomarkers. This biofabrication platform provides a scalable framework for drug screening and biomarker discovery, with immediate applications in precision medicine for patient stratification and combination therapy development.

cancer biology↗

bioRxiv2025

Targeted degradation of SARS-CoV-2 via the autophagy-lysosome system using chemical mimetics of the N-degron pathway

In the N-degron pathway, ATE1 transfers the amino acid L-arginine (L-Arg) from Arg-tRNAArg to N-terminal (Nt) residues of cellular proteins. The resulting Arg/N-degrons bind the autophagic receptor p62/SQSTSM-1/Sequestosome-1 to induce lysosomal degradation of various biomaterials. Here, we demonstrate that the chemical mimetics of Arg/N-degrons, termed autophagy-targeting ligands (ATLs), can induce lysosomal degradation of SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) via p62-mediated macroautophagy. In Vero E6 cells infected with SARS-CoV-2, ATLs promoted p62 self-polymerization and enhanced LC3 synthesis and lipidation, leading to viral sequestration within autophagosomes for lysosomal degradation. In transgenic mice overexpressing human angiotensin-converting enzyme 2 (ACE2), oral administration of ATL1014 inhibited viral replication and increased viability. In a Syrian hamster model, ATL1014 attenuated viral replication in the lungs and demonstrated efficacy in inflammatory lesions and pulmonary congestions. These results identify the N-degron pathway as a potential target for a host-targeting strategy (HTS) against a broad spectrum of viruses.

microbiology↗

bioRxiv2025

MIA-Jet: Multi-scale Identification Algorithm of Chromatin Jets

The mammalian genome is organized into large-scale chromosome territories, compartments, domains, and at the smallest scale, chromatin loops and stripes. The newest element is a chromatin jet, a diffused line perpendicular to the main diagonal in the Hi-C contact map, which was reported in quiescent mammalian lymphocytes supporting a two-sided symmetric cohesin loop extrusion model. A similar structure is observed in Repli-HiC and related data, where relatively thin and straight chromatin fountains indicate coupling of DNA replication forks. However, the precise biological implications of these jet-like structures are unknown due to the limitations in computational methods. We developed MIA-Jet, a multi-scale ridge detection algorithm that can accurately detect jets of variable lengths, widths, and angles. When tested on Hi-C, Repli-HiC, ChIA-PET, ChIA-Drop, and Micro-C data in mouse, human, roundworm, and zebrafish cells, MIA-Jet outperformed existing methods. In human cells, jets were enriched in cohesin loading sites and early replication initiation zones. Applying MIA-Jet to Hi-C data generated from protein-degraded cells revealed that jets are dependent on cohesin and that depleting CTCF results in longer and less angled jets than wild-type. We envision MIA-Jet to be broadly applicable to any 3D genome mapping data, thereby providing new insights into the functional roles of chromatin jets.

bioinformatics↗

bioRxiv2025

Executive control in Obsessive-Compulsive Disorder: A worldwide mega-analysis of task-based functional neuroimaging data of the ENIGMA-OCD consortium

ObjectiveObsessive-compulsive disorder (OCD) is associated with impaired executive function and altered activity in associated neural circuits, contributing to reduced goal-directed behavior. To investigate neural activation during executive control, we conducted a mega-analysis in the ENIGMA-OCD consortium pooling individual participant data from 475 individuals with OCD and 345 healthy controls across 15 fMRI tasks collected worldwide. MethodsIndividual participant data was uniformly processed using HALFpipe to construct voxelwise statistical images of executive control and task load contrasts. Parameter estimates extracted from regions of interest were entered into multilevel Bayesian models to examine regional and whole-brain effects of diagnosis, and, within OCD, the influence of medication status, symptom severity, and age of onset on task activation. ResultsWe observed a robust task activation pattern across individuals with OCD and control participants in executive control regions across tasks. Relative to controls, individuals with OCD showed moderate to very strong evidence of weaker activation of the dorsolateral prefrontal cortex, precuneus, frontal eye fields, and inferior parietal lobule during executive control (all positive posterior probabilities [P+]<0.1). Individuals with OCD also showed stronger activation in regions of the default mode network during executive function relative to controls. We found little evidence for differential activation during executive control in task- positive regions related to disease onset, severity and medication status. ConclusionIn the first mega-analysis of fMRI studies of executive function in OCD, we found strong evidence of weaker frontoparietal activation during executive control tasks. Our findings also suggest a failure of default mode network regions to appropriately disengage during task performance in OCD.

neuroscience↗

bioRxiv2025

SALL4 is required for YAP1-dependent malignant and regenerative hepatocyte reprogramming into cholangiocyte lineage

Hepatocytes (HCs), which share a developmental origin with cholangiocytes (CCs), have the capacity to undergo reparative reprogramming into CCs in response to liver injury and, under specific conditions, can also transform malignantly into cholangiocarcinoma (CCA). However, the molecular mechanisms governing HC plasticity in liver diseases remain poorly understood. In this study, we investigated the role of Spalt Like Transcription Factor 4 (SALL4), an oncofetal transcription factor, in both malignant and regenerative HC fate transitions toward the biliary lineage. Using Sleeping Beauty hydrodynamic tail vein injection-mediated murine liver cancer models, we explored HC-to-CCA transformation, while the DDC diet-induced cholestasis model was used to investigate regenerative HC-to-CC reprogramming. Our findings reveal that SALL4 is specifically required for myristoylated Akt (myrAkt)-YAP1S127A (AY)-driven HC-to-CCA transformation, as its loss significantly suppressed malignant reprogramming and clonal expansion. Surprisingly, SALL4 overexpression also prevented AY-driven CCA development while promoting the expansion of liver progenitor cell (LPC)-like fatty HCs. Mechanistically, we propose Bmi1 as a key downstream effector of SALL4 in YAP1-dependent HC- to-CCA transformation. Additionally, in the DDC-fed cholestasis model, Sall4 deletion enhanced HC-to-LPC activation while impairing LPC differentiation into mature CCs. These findings establish SALL4 as a critical regulator of HC plasticity in both malignant and regenerative contexts and highlight its potential as a therapeutic target for specific liver cancer subtypes. SIGNIFICANCEHepatocyte plasticity supports repair but can drive malignancy, acting as a double-edged sword. We identify SALL4 as regulator of YAP1-driven hepatocyte-to-cholangiocyte reprogramming, revealing the YAP1-SALL4- BMI1 axis as a therapeutic target for cholangiocarcinoma.

cancer biology↗

bioRxiv2025

Chrom-Sig: de-noising 1-dimensional genomic profiles by signal processing methods

MotivationModern genomic research is driven by next-generation sequencing experiments such as ChIP-seq, CUT&Tag, and CUT&RUN that generate coverage files for transcription factor binding, as well as ATAC-seq that yield coverage files for chromatin accessibility. Due to the inherent technical noise present in the experimental protocols, researchers need statistically rigorous and computational efficient methods to extract true biological signal from a mixture of signal and noise. However, existing approaches are often computationally demanding or require input or spike-in controls. ResultsWe developed Chrom-Sig, a Python package to quickly de-noise 1-dimensional genomic coverage tracks by computing the empirical null distribution without prior assumptions or experimental controls. When tested on 19 ChIP-seq, CUT&RUN, ATAC-seq, snATAC-seq datasets, Chrom-Sig can effectively decompose the data into signal and noise. Notably, Chrom-Sig performs de-noising and peak calling in 1-2 hours using around 20 GB of memory. The de-noised signal corroborates with biologically meaningful results: CTCF CUT&RUN data retained a higher percentage of peaks overlapping CTCF binding motifs and ATAC-seq and RNA Polymerase II data resulted in enhancers and promoters. We envision Chrom-Sig to be a highly versatile and general tool for current and future genomic technologies. AvailabilityChrom-Sig is publicly available at https://github.com/minjikimlab/chromsig under the MIT license. Contactminjilab@umich.edu

bioinformatics↗

bioRxiv2025

LPS O-antigen polysaccharide length impacts outer membrane permeability of enteric gram-negative bacteria

The Gram-negative outer membrane (OM) forms the bacterial cell surface and acts as a barrier against antibiotic influx. In enteric species, the OM is covered by lipopolysaccharides (LPS) decorated with varying lengths of O-antigen (O-Ag) polysaccharide that protect bacteria against mammalian host defenses. Studies of lab-adapted Escherichia coli K-12 strains have proven instrumental in unravelling the essential processes of LPS synthesis, transport, and assembly into the OM. However, O-Ag synthesis was inactivated in K-12 strains during their lab adaption, and these cells produce a non-native, truncated LPS form. Surprisingly, we found that re-activating O-Ag synthesis in K-12 permeabilizes the OM to diverse antibiotics, causing susceptibility. The O-Ag that modifies LPS is directly responsible for the compromised OM barrier. Lengthening the O-Ag polysaccharide worsens antibiotic sensitivity; while shortening it, or removing it entirely, improves antibiotic resistance in both E. coli and the human pathogen, Shigella flexneri. Our data show that OM antibiotic barrier integrity is maintaining by a balanced production of long and short LPS forms, and that this balance is dysfunctional in model E. coli K-12 strains. Our findings reveal that long O-Ag polysaccharides are a double-edged sword: while well-recognized as critical for protection against external host assaults, their transport and assembly onto the surface comes at the inherent price of compromising the OM barrier. Hence, LPS production balances between competing needs in host defense and OM integrity. Moreover, we identify an inherent advantage for species that produce O-Ag-lacking lipooligosaccharide (LOS), rather than LPS. IMPORTANCEThe outer membrane (OM) of bacteria like Escherichia coli and Shigella flexneri forms a barrier that protects cells against antibiotics and immune effectors. The surface-exposed leaflet is filled by lipopolysaccharides (LPS) decorated with long "O-antigen" (O-Ag) polysaccharides. The benefit of covering the surface with O-Ag is well-appreciated - these long polysaccharides shield against host assaults. Our study reveals a hidden cost to these long O-Ag polysaccharides: transporting and assembling LPS modified with O-Ag compromises integrity of the OM antibiotic barrier, rendering bacteria vulnerable to antibiotics. Cells must balance O-Ag across two parameters - protection from the host and preserving OM integrity. Our findings also present an inherent benefit to not producing O-Ag, a common feature among diverse bacterial pathogens.

microbiology↗

bioRxiv2025

Proteostasis and Unfolded Protein Response Dynamics in Human Neuron and Mouse Glia Co-culture Reveal Cell-Specific Aging Responses

Proteostasis, or protein homeostasis, is a tightly regulated network of cellular pathways essential for maintaining proper protein folding, trafficking, and degradation. Neurons are particularly vulnerable to proteostasis collapse due to their post-mitotic and long-lived nature and thus represent a unique cell type to understand the dynamics of proteostasis throughout development, maturation, and aging. Here, we utilized a dual-species co-culture model of human excitatory neurons and mouse glia to investigate cell type- specific, age-related changes in the proteostasis network using data-independent acquisition (DIA) LC-MS/MS proteomics. We quantified branch-specific unfolded protein response (UPR) activation by monitoring curated effector proteins downstream of the ATF6, IRE1/XBP1s, and PERK pathways, enabling a comprehensive, unbiased evaluation of UPR dynamics during neuronal aging. Species-specific analysis revealed that aging neurons largely preserved proteostasis, although they showed some signs of collapse, primarily in ER-to-Golgi transport mechanisms. However, these changes were accompanied by upregulation of proteostasis-related machinery and activation of the ATF6 branch, as well as maintenance of the XBP1s and PERK branches of the UPR with age. In contrast, glia exhibited broad downregulation of proteostasis factors and UPR components, independent of neuronal presence. Furthermore, we quantified stimulus-specific modulation of select UPR branches in aged neurons exposed to pharmacologic ER stressors. These findings highlight distinct, cell-type-specific stress adaptations during aging and provide a valuable proteomic resource for dissecting proteostasis and UPR regulation in the aging brain. SignificanceUnderstanding how the unfolded protein response (UPR) and proteostasis network change with age is often studied in model organisms, where pathways are assessed across mixed cell types. Such systems can obscure cell-type-specific regulation. Here, we evaluate age-associated remodeling of the UPR and proteostasis network in a dual-species co-culture of human neurons and mouse glia using DIA proteomics. This approach enables species-specific proteomic profiling without physical separation, supported by a customizable data analysis pipeline. We show that neurons and glia exhibit divergent age-related responses, with neurons maintaining adaptive proteostasis and glia showing broader declines. The analytical framework presented here supports future studies to uncover additional cell-type-specific aging phenotypes or to probe the effects of pharmacologic or physical manipulation of biological systems.

biochemistry↗

bioRxiv2025

Combinatorial action of regulatory systems generates colistin heteroresistance

Heteroresistance is a form of antibiotic resistance in which a minor subpopulation of resistant cells coexists with a majority susceptible population. Colistin heteroresistance is common among Enterobacter cloacae clinical isolates, threatens its utility as a last-line therapeutic, and has become a model with which to understand the fundamental bases of heteroresistance. Despite numerous insights, the mechanism by which phenotypic heterogeneity is generated within the population and leads to colistin heteroresistance has been unclear. Here, using a transposon-based mutagenesis screen, we identify the sigma factor {sigma}E as the source of heterogeneity in population-wide colistin resistance levels. Single-cell tracking experiments revealed that {sigma}E is active in only one percent of the population at baseline, and only those cells with active {sigma}E survive colistin exposure. However, {sigma}E expression and population heterogeneity are insufficient for survival, as a mutant lacking the PhoPQ two-component system controlling lipid A modifications necessary for colistin resistance retains heterogeneity but loses colistin resistance. These findings lead to a new paradigm in heteroresistance, where the combinatorial action of multiple regulatory systems, encompassing a heterogeneity generator and a distinct resistance generator, are required to give rise to colistin heteroresistance.

microbiology↗

bioRxiv2025

High-Resolution Detection of Hidden Antibiotic Resistance with the Dilution-and-Delay (DnD) Susceptibility Assay

Rising rates of antibiotic treatment failure highlight the complexity of resistance mechanisms. While conventional, genetically encoded resistance is well established, recent studies have uncovered widespread non-canonical mechanisms driven by phenotypically insensitive subpopulations hiding in seemingly susceptible populations, such as heteroresistance, persistence or adaptive resistance. These variants are not only clinically significant but also act as evolutionary precursors to fully resistant populations, rendering infections increasingly difficult to control. Yet standard antibiotic susceptibility tests lack the resolution and dynamic range needed to detect this wide spectrum of resistance mechanisms and their evolutionary progression, often misguiding clinical decisions, obscuring mechanisms of failure, and underestimating the true epidemiology of antibiotic resistance. To address these limitations, we introduce the Dilution-and-Delay (DnD) assay--a practical, high-resolution approach that implements two basic principles of bacterial growth in the antibiotic susceptibility testing format. Using well-defined synthetic communities and strains, we demonstrate that the DnD assay quantifies viable cells over more than eight orders of magnitude and detects rare antibiotic-insensitive cells at frequencies as low as 1 in 100 million. It additionally reports the standardized MIC as a secondary output, thereby capturing the average antibiotic response of the majority population and heterogeneity of minority subpopulations within a single assay. We scaled this approach for high-throughput application, classifying [~]120 previously uncharacterized clinical isolates of Klebsiella pneumoniae, Enterobacter cloacae, Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumannii--across five antibiotics. This robust, quantitative, and scalable platform opens the door to next-generation antibiotic susceptibility testing, with broad utility in basic research, clinical diagnostics, and epidemiological surveillance. This improved testing will guide evidence-based clinical decisions and prevent the rapid evolution of antibiotic resistance, thereby counteracting the rising rate of treatment failure. SignificanceAntibiotic treatment failure due to inappropriate prescription remains a critical threat to global health. It also accelerates the evolution of resistance, making infection increasingly difficult to control. A key underlying issue is that current antibiotic susceptibility testing lacks the resolution and dynamic range needed to detect a wide range of resistance mechanisms and their evolutionary progression, leading to misinformed clinical decisions. While population analysis profiling (PAP) offers sufficient accuracy, it is too labor-intensive for routine use. Single-cell approaches are technically demanding and resource-intensive. Here, we present a susceptibility testing strategy that preserves the operational efficiency and compatibility of conventional workflows, while dramatically extending their resolution and dynamic range. Our approach is scalable, quantitative, and well-suited for high-throughput applications in both research and clinical settings--closing a critical diagnostic gap in the fight against antibiotic treatment failure.

microbiology↗

bioRxiv2025

Beyond Gatekeeping: Efflux Pumps Remotely Destabilize Cytoplasmic Drug-Target Interactions by Limiting Rebinding.

Bacterial efflux pumps are major contributors to multidrug resistance, classically described as "gatekeepers" that reduce drug entry. Here, we uncover an equally potent post-entry mechanism of efflux pumps, revealing that their function extends deep into intracellular drug-target interactions. Using quantitative live-cell fluorescence imaging, we monitored the activity of major efflux systems in Escherichia coli and Pseudomonas aeruginosa with Hoechst (HCT), a DNA-binding inhibitor. As expected, more HCT entered efflux-deficient cells than efflux-active cells. Unexpectedly, the intracellular HCT in efflux-deficient cells also bound DNA more stably, resulting in higher binding affinity. Statistical-physics-based modeling and experimental testing show that that unlike in dilute in vitro conditions, drug molecules that unbind from their targets in intracellular environments rapidly undergo successive rebinding, prolonging the lifetime of the drug-target complex. However, efflux pumps counteract this effect by suppressing rebinding, thereby kinetically destabilizing drug-target interactions and lowering the binding affinity. This kinetic destabilization acts synergistically with canonical gatekeeping to broaden and amplify drug resistance. Our findings reveal an underappreciated biophysical mechanism of efflux pumps, expanding the classical biochemical view of how efflux pumps confer multidrug resistance.

microbiology↗

bioRxiv2025

Distributed control circuits across a brain-and-cord connectome

Just as genomes revolutionized molecular genetics, connectomes (maps of neurons and synapses) are transforming neuroscience. To date, the only species with complete connectomes are worms1-3 and sea squirts4 (103-104 synapses). By contrast, the fruit fly is more complex (108 synaptic connections), with a brain that supports learning and spatial memory5,6 and an intricate ventral nerve cord analogous to the vertebrate spinal cord7-11. Here we report the first densely reconstructed adult fly connectome that unites the brain and ventral nerve cord, and we leverage this resource to investigate principles of neural control. We show that effector neurons (motor neurons, endocrine cells and efferent neurons targeting the viscera) are primarily influenced by sensory neurons in the same body part, forming local feedback loops. These local loops are linked by long-range circuits involving ascending and descending neurons organized into behavior-centric modules. Single ascending and descending neurons are often positioned to influence the voluntary movements of multiple body parts, together with the endocrine cells or visceral organs that support those movements. Brain regions involved in learning and navigation supervise these circuits. These results reveal an architecture that is distributed, parallelized and embodied, reminiscent of distributed control architectures in engineered systems12,13.

neuroscience↗

bioRxiv2025

Structural Variants at Intronic CTCF Loop Anchors Drive Differential Exon Usage

CTCF-mediated chromatin loops are known to influence gene regulation, yet their role in pre-mRNA splicing remains incompletely understood. Here, we demonstrate that structural variants (SVs) at the anchors of intronic CTCF loops can modulate exon usage. By integrating high-resolution three-dimensional (3D) genome organization and gene expression datasets from C57BL/6J (B6) and 129S1/SvImJ (129S) mouse embryonic stem cells (ESCs), with structural variant (SV) maps from the 129S mouse, we identified thousands of intron-anchored CTCF loops. Our data indicate that SVs intersecting loop anchors are more frequently associated with differential exon inclusion events than with changes in overall gene expression. CRISPR/Cas9 deletion of two SV-harboring intronic CTCF sites in Numbl and Ireb2 validated the predicted splicing shifts observed between B6 and 129S ESC that correspond with diminished long-range chromatin looping. Our findings reveal a direct mechanistic link between 3D genome architecture and alternative splicing and highlight non-coding SVs as modulators of transcript diversity. Our study has thus identified a novel class of CTCF-bound regulatory elements regulating alternative splicing. Cataloging and validating these functional elements will elucidate molecular mechanisms underlying phenotypic variation within populations.

molecular biology↗

bioRxiv2025

Turmeric Phyto-nanoparticle: enhanced cellular bioavailability and anti-inflammatory effect in human monocyte / macrophage model

The poor bioavailability of curcuminoids remains a major challenge to therapeutic use. This is largely due to their hydrophobicity, poor absorption, rapid metabolism, and short circulating half-life--limitations that are now being addressed through advances in nano- and micro-emulsion technologies. Curcuminoids and other water-insoluble phyto-polyphenols offer significant putative health benefits as anti-inflammatory, antioxidant, anticancer, radioprotective, and neuroprotective agents. Conventional emulsion-based delivery systems, such as liposomes, micelles, or solid lipid particles, rely on various emulsifying surfactants and/or excipients, some of which may themselves pose health risks. Here, we establish a novel class of all-natural, additive-free, oil-free, and emulsion-free Turmeric Phyto-NanoParticles (TPNPs) formulated directly from turmeric rhizomes and tested in a human monocyte/macrophage cell model to assess bioavailability kinetics and the efficacy of antioxidant and anti-inflammatory potential. TPNPs are enriched with curcuminoids (24.85% by mass), form a homogeneous nanoparticle distribution, exhibit higher antioxidant capacity, and demonstrate significantly improved cellular uptake in both monocytes and macrophages compared to conventionally purified curcuminoids. Favourable cellular pharmacodynamic anti-inflammatory effect of TPNPs was shown by increased levels of the cytoprotective enzyme heme oxygenase-1 (HMOX1), and a more effective reduction in lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF) secretion compared to conventional curcuminoids. TPNPs could thus serve as a stable, non-synthetic, excipient-free formulation for safe and effective delivery of curcuminoids by nanocarriers for inflammatory conditions.

pharmacology and toxicology↗

bioRxiv2025

Polycomb repression works without Siesta

Polycomb group proteins mediate epigenetic repression via multi-subunit complexes, including canonical Polycomb Repressive Complex 1 (PRC1), which monoubiquitylates histone H2A and binds histone H3 tri-methylated at lysine 27 (H3K27me3). The RING1 subunit of PRC1, critical for H2A ubiquitylation, forms other complexes. These variant RING1 complexes also ubiquitylate H2A but cannot bind H3K27me3, and their evolutionary origins and functions are debated. Using Drosophila genetics, we discovered that canonical PRC1 and variant RING1 complexes ubiquitylate H2A at distinct genomic regions. We established that the sole Drosophila PCGF protein specific for variant RING1 complexes, which we named Siesta, is not required for epigenetic repression of developmental genes, but controls larval locomotion independently of H2A ubiquitylation. Leveraging a massively parallel transgenic approach, we demonstrated that H2A ubiquitylation has minimal impact on transcriptional repression. Our findings imply that variant RING1 complexes operate outside the Polycomb regulatory system and that the popular PRC1 nomenclature needs revision.

genetics↗

bioRxiv2025

Plagl1 and Lrrc58 control mammalian body size by triggering target-directed microRNA degradation of miR-322 and miR-503

Precise control of microRNA (miRNA) expression is critical during development. An important mechanism of miRNA regulation is target-directed microRNA degradation (TDMD), a pathway in which the binding of miRNAs to specialized trigger RNAs induces ubiquitylation and decay of associated Argonaute (AGO) proteins by the ZSWIM8 ubiquitin ligase. Concomitant release of miRNAs results in their rapid turnover. ZSWIM8-deficient mice exhibit reduced body size, cardiopulmonary and neurodevelopmental defects, and perinatal lethality. Despite widespread dysregulation of miRNAs in these animals, the vast majority of presumptive trigger RNAs that induce decay of ZSWIM8-regulated miRNAs remain undefined. Here, using AGO crosslinking and sequencing of hybrids (AGO-CLASH), a high-throughput method for identifying miRNA binding sites, we report the identification of Plagl1 as a TDMD trigger for miR-322-5p, and Lrrc58 and Malat1 as TDMD triggers for miR-503-5p in mouse embryonic fibroblasts (MEFs). In mice, deletion of the miR-322-5p and miR-503-5p trigger sites in the Plagl1 and Lrrc58 3' UTRs, respectively, abrogated TDMD of these miRNAs and resulted in miR-322/503-dependent embryonic growth restriction, recapitulating a key feature of the Zswim8--/-- phenotype. Thus, Plagl1 and Lrrc58 act as triggers for degradation of miR-322-5p and miR-503-5p, revealing a noncoding function for these mRNAs as regulators of mammalian body size.

molecular biology↗

bioRxiv2025

Untargeted Plasma Metabolomics in Canine Cognitive Dysfunction: The Naturally Occurring Alzheimer's Disease Analog in Dogs

IntroductionCanine Cognitive Dysfunction (CCD) is an increasingly prevalent naturally occurring neurodegenerative condition in senescent dogs that share neuropathological and clinical features with human Alzheimers disease (AD). Metabolic profiling allows for identification of new candidates for AD biomarkers, diagnostics, and therapeutics. Despite its translational potential, plasma metabolomic profiling of dogs with CDD has not been previously characterized. MethodsThis case-control study analyzed plasma samples from ten client-owned geriatric dogs, including five with severe CCD and five age-matched, clinically healthy controls. Untargeted plasma metabolomics was performed using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). Multivariate and univariate statistical analyses identified significant metabolic differences between the groups. Metabolites were considered significant based on a variable importance in projection (VIP) score > 1.5, fold change (FC) > 2.0, and adjusted p-value < 0.05. ResultsFifteen metabolites across seven chemical classes were significantly altered in CCD dogs compared to controls, including glycerophospholipids, steroid derivatives, indoles, and mitochondrial-related compounds. Notably, elevated lysophosphatidic acid (LPA 20:2/0:0) and reduced ubiquinone-2 levels suggest dysregulation in neuroinflammatory and oxidative stress pathways. Cholesterol exhibited the highest FC and VIP scores, further reinforcing its role in AD pathogenesis. Hierarchical clustering and pathway enrichment analyses supported distinct metabolic signatures in CCD that mirror those observed in human AD. DiscussionThis is the first untargeted plasma metabolomic profiling of dogs with CCD, revealing systemic metabolic disturbances that align with AD pathophysiology. Data was collected from senescent community-dwelling companion dogs, which enhances the studys ecological and translational relevance. It supports the utility of CCD as an AD model and highlight candidate plasma biomarkers that warrant further investigation. Future longitudinal studies integrating metabolomics with neuroimaging, histopathology, and behavioral assessments are required to validate these findings and contribute to AD biomarker discovery and therapeutic development.

neuroscience↗

bioRxiv2025

Murine peritoneal macrophages undergo female-specific remodeling with aging

Aging is a complex process characterized by a progressive decline in physiological functions. Immune function is strongly influenced by biological sex, affecting both innate and adaptive responses. Here, we investigated the effects of age and sex on mouse peritoneal immune cells and identified macrophages as the top affected cell type. Macrophages, as central components of the innate immune system, play critical roles in homeostasis maintenance and infection response. We found that aging induces sex-specific remodeling of mouse peritoneal macrophage omic landscapes, which is accompanied by female-specific age-related functional remodeling (i.e. decreased phagocysis, increased glycolysis). We show that age-related changes in circulating estrogen levels likely drive aspects of female-specific macrophage age-related changes, specifically age-related phagocytosis decline. By leveraging our multi-omic dataset, we identify transcription factors whose female-specific age-regulation may drive female-specific age-related changes in peritoneal macrophage phenotypes. Interestingly, Irf2 downregulation was sufficient to recapitulate aspects of female peritoneal macrophage aging phenotypes, including increased glycolysis. Mechanistically, decreased Irf2 expression leads to decreased binding to its target genes, including Hk3, which encodes hexokinase (the rate-limiting enzyme of glycolysis). Our findings support the notion that aging leads to female-specific remodeling of mouse peritoneal macrophages through hormone-dependent and -independent mechanisms.

immunology↗