Biology subjects

Shin, S.

Publications and source records attributed to Shin, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Human caspase-4 and caspase-5 assemble into non-canonical inflammasomes in response to overlapping and distinct LPS structures

Innate immune sensing of bacterial lipopolysaccharide (LPS) is critical to host defense, but a dysregulated response can lead to sepsis, a life-threatening condition associated with multiple organ failure and shock. LPS is sensed at the cell surface by Toll-Like Receptor 4 (TLR4) and in the cytosol by mouse caspase-11 (Casp11) or its human orthologs, caspase-4 and 5 (CASP4/5). In response to cytosolic LPS, Casp11 oligomerizes in a catalytically dependent manner into supramolecular organizing centers (SMOCs). We demonstrate that CASP4 and CASP5 also form catalytically dependent SMOCs in response to bacterial LPS in macrophages. Furthermore, we find that CASP4 and CASP5 differentially respond to distinct LPS structural variants that fail to activate CASP11 and that only activating LPS structures induce SMOC formation by the corresponding caspase. Notably, transgenic mice expressing CASP4 in lieu of Casp11 exert more effective control of Francisella novicida, a rodent pathogen whose LPS evades TLR4 and Casp11 detection. Conversely, CASP4-expressing mice are more susceptible to sepsis when exposed to E. coli expressing a lipid A variant that fails to activate TLR4 and Casp11. Together, these findings reveal human CASP4 and CASP5 have evolved to assemble into SMOCs in response to a broader range of LPS structures than mouse Casp11 and highlight the potential tradeoff between systemic inflammation and control of bacterial infection that may underline these differences. Our findings may inform the development of targeted therapeutic strategies for gram-negative bacterial infection and sepsis that are effective in humans.

immunology↗

bioRxiv2026

Extracellular cGAMP transmission exacerbates neuroinflammation in aging and multiple sclerosis

Neuroinflammation drives neurodegeneration and cognitive decline in aging, but the signals that spread this inflammation across the CNS remain unclear. Here, we show in mice that advanced age and experimental autoimmune encephalomyelitis (EAE) both result in elevated cytosolic mtDNA in microglia, which stimulates the production and export of cGAMP from these cells. Meanwhile, ENPP1, the dominant cGAMP hydrolase in the brain and spinal cord, is downregulated in both EAE mice and MS patients, releasing the brake on extracellular cGAMP accumulation. In Enpp1H362A mice, which selectively lack cGAMP hydrolysis activity, unrestrained extracellular cGAMP worsens EAE-associated paralysis and drives premature aging phenotypes including impaired motor coordination (rotarod, pole test) and reduced exploration (open field test) in a manner dependent on STING. Single-cell RNA sequencing pinpoints microglia, astrocytes, and inhibitory neurons as the principal cGAMP responders that activate STING-dependent interferon-stimulated gene induction in response to elevated extracellular cGAMP. Mechanistically, microglia import cGAMP through the LRRC8A:C channel, while astrocytes and neurons import cGAMP via the glutamate antiporter system xc-, reinforced by SLC1A2-mediated glutamate uptake in astrocytes and SLC38A1/2-fueled glutamine-to-glutamate synthesis in neurons. In isolated primary astrocytes and neurons, high intracellular glutamate and system xc--driven redox demands favor cGAMP uptake, raising the possibility that neurotransmitter cycling and redox burden shape cellular vulnerability to cGAMP immunotransmission in vivo. Together, these findings position ENPP1 as a protective brake on neuroinflammation in aging and MS, nominating cGAS inhibition and CNS ENPP1 enhancement as therapeutic strategies.

cell biology↗

bioRxiv2026

The polysaccharide portion of LPS is critical for caspase-11 inflammasome activation

Caspase-11 (Casp11) is a murine zymogen that becomes active upon sensing cytosolic lipopolysaccharide (LPS), enabling it to form the non-canonical inflammasome and execute an inflammatory cell death known as pyroptosis. The lipid A acylation state of LPS is thought to be the primary regulator of Casp11 activation. However, our data demonstrate that the polysaccharide moiety of LPS, and particularly the polysaccharide phosphates, are critical for eliciting maximal Casp11 activation. Accordingly, the polysaccharide was required for Rhodobacter sphaeroides LPS to act as an inflammasome antagonist. Furthermore, isolated polysaccharide was sufficient to antagonize Casp11 activation. Mechanistically, our findings support a two-step model of non-canonical inflammasome activation where the polysaccharide recruits Casp11 to LPS and the lipid A acylation state governs oligomerization efficiency. These findings provide new insight into the molecular basis of LPS-driven Casp11 activation, with implications for the design of future therapies that target the non-canonical inflammasome.

microbiology↗

bioRxiv2026

Distinct kinetics and mechanisms of microbial inactivation of enteric virus revealed by capsid and genome integrity

Enteric viruses are important contaminants of surface waters and a significant burden on public health. In aquatic ecosystems, the stability of these pathogens is differentially impacted by abiotic and biotic stressors, which can exert inactivating effects. Here, we investigated the fate of two enteroviruses, echovirus 11 (E11) and coxsackievirus B5 (CVB5), and one adenovirus, human adenovirus 2 (HAdV2), in lakewater and explored the mechanisms underlying their microbial inactivation. By combining infectivity assays, genome quantification, and capsid integrity analysis, we characterized virus-specific inactivation kinetics and mechanisms and examined the relationship between infectivity loss and capsid structural integrity. We observed rapid, intermediate, and negligible decay for HAdV2, E11, and CVB5, respectively, and confirmed that microbial proteases contribute to their inactivation. In addition, we revealed that loss of capsid structural integrity drives E11 inactivation, but not HAdV2 inactivation. Genome decay did not consistently correlate with loss of infectivity, highlighting the limitations of genome-based detection for assessing the presence of infectious viruses. These findings provide new insights into the mechanisms governing virus inactivation in aquatic environments and emphasize the importance of understanding viral fate when interpreting molecular detection data to assess virus-associated microbial risks.

microbiology↗

bioRxiv2026

Redox activation of ATG5 licenses autophagy upon nutrient restriction

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

cell biology↗

bioRxiv2026

A quantitative redox proteome of the human muscle response to exercise

Reactive oxygen species (ROS) regulate protein function through reversible cysteine oxidation. In human skeletal muscle, exercise-induced ROS initiates adaptations such as mitochondrial biogenesis, increased insulin sensitivity, and hypertrophy. However, specific protein targets of ROS regulation during exercise remain unclear owing to longstanding challenges in analyzing redox proteomes in vivo. We applied cysteine derivatization and multiplexed proteomics to map muscle protein cysteine oxidation in humans during exercise. The OxiMuscle dataset quantifies reversible modifications across 9,177 unique cysteine sites on 2,782 proteins, comprising 17,492 individual cysteine site measurements in young men undergoing three types of exercise, providing the first comprehensive, site-resolved and quantitative analysis of the exercise-regulated redox cysteine proteome in humans. We systematically define cysteine oxidation targets regulated by at least one form of exercise, many of which reside in proteins with established roles in muscle physiology. Among these sites is a redox-regulated cysteine on the autophagy receptor protein p62. We demonstrate that reversible oxidation of this cysteine regulates p62-mediated autophagy upon myotube contraction and mouse muscle adaptation to exercise in vivo. Together, these results define a redox-driven mechanism linking exercise-induced autophagy to muscle adaptation. More broadly, our findings offer a comprehensive resource on redox-signaling networks in human muscle, accessible at http://oximuscle-alb-1899330623.us-east-1.elb.amazonaws.com/.

cell biology↗

bioRxiv2026

Holotomography reveals biophysical remodeling of mouse oocytes during post-ovulatory aging

Routine assessment of oocyte state in reproductive biology and clinical embryology still relies largely on two-dimensional transmitted-light morphology, leaving the underlying three-dimensional intracellular organization largely unmeasured. Here, we use holotomography to reconstruct volumetric refractive index (RI) distributions in unlabeled live mouse oocytes and combine this imaging with sparse-annotation, AI-assisted compartment segmentation and multidimensional feature extraction across geometry, RI statistics, dry-mass density, and intracellular texture. Applied to an in vitro model of post-ovulatory aging, holotomography-derived biophysical profiles separated fresh and aged oocytes more strongly than two-dimensional brightfield-derived features, and feature-family ablation indicated that non-morphological descriptors contributed substantially to this separation. At the population level, aged oocytes showed coordinated ooplasmic remodeling, including reduced volume, increased RI heterogeneity, and increased dry-mass density while total dry mass was broadly conserved. These findings demonstrate a label-free quantitative framework for detecting post-ovulatory aging-associated biophysical remodeling in intact mammalian oocytes. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=167 SRC="FIGDIR/small/733271v1_ufig1.gif" ALT="Figure 1"> View larger version (89K): org.highwire.dtl.DTLVardef@c7257aorg.highwire.dtl.DTLVardef@6d92f0org.highwire.dtl.DTLVardef@1c9f7eborg.highwire.dtl.DTLVardef@b36637_HPS_FORMAT_FIGEXP M_FIG C_FIG

Developmental Biology↗

bioRxiv2026

Self-feeding living materials enabled by cell responsive glycogen nanoparticles as metabolic batteries

Scaling engineered living materials to clinically relevant dimensions is limited by diffusion-dependent depletion of oxygen and nutrients, which rapidly induces metabolic failure. We introduce glycogen as nutritional nanoparticle that provides cell-mediated, autonomous nutrient release to support long-term survival under extreme metabolic stress. We demonstrate that human mesenchymal stromal cells (hMSCs) survive for weeks in anoxia and serum deprivation when provided extracellular glycogen. Contrary to long-held assumptions, hMSCs secrete glycogen-degrading enzymes, enabling cell-density controlled extracellular glycogenolysis and sustained release of glucose and metabolic intermediates, positioning glycogen as the first-of-its-kind metabolic battery. This cell-responsive process maintains metabolic activity, limits glycolytic acidosis, and enhances pro-angiogenic signaling. To translate this mechanism into a versatile materials platform, we engineered core-shell dextran-tyramine microcapsules that stably encapsulate glycogen while permitting diffusion of enzymes and degradation products. Integrated into centimeter-scale GelMA constructs, these microcapsules maintained hMSC viability and function for at least one month under anoxia. In vivo, glycogen-loaded implants promote deep cellular infiltration, enhanced matrix remodeling, increased M2 macrophage polarization, and orchestrated accelerated vascularization. This work establishes the novel concept of glycogen-based nutritional nanoparticles as metabolic batteries to endow engineered tissues with autonomous self-feeding capacity, enabling scalable and functional living materials for regenerative medicine and related technologies.

bioengineering↗

bioRxiv2026

The Culicinae are Monophyletic and Ancient: A response to Pierce et al. 2025

Mosquitoes are classified into two subfamilies, each monophyletic, and typically considered to both be ancient, having diverged more than 100 million years ago based on previous divergence analyses. A recent publication challenged this view with phylogenomic results primarily from the third codon position and UCEs. Utilizing alternative fossil placement and these phylogenomic data, these authors find that the Culicidae and Chaoboridae diverged in the lower Cretaceous, and that one mosquito subfamily, the Anophelinae, is nested within the Culicinae. These results are in stark contrast to previous results from diverse data sources, ranging from other genomic data, to morphology, to fossils. Here, we briefly detail the substantial evidence that supports two monophyletic subfamilies of extant mosquitoes, along with fossil evidence that supports the ancient divergence of these lineages.

evolutionary biology↗

bioRxiv2026

Connecting multiway enhancer-promoter interactions to changes in gene expression in cancer

Hi-C, and more recently Micro-C, experiments suggest that genome organization is altered as normal cells become cancerous, often resulting in aberrant gene expression. In prostate cancer, there is evidence that large topologically associating domains in normal cells split and are accompanied by shifts in the epigenetic marks from inactive to active states. However, it is unclear whether changes in genome organization alone can account for the gene expression increase in cancer cells. By combining polymer physics concepts and data-driven modeling, we first calculated an ensemble of three-dimensional (3D) chromatin structures using only the 2D contact map as input. The enhancer-promoter (E-P) distance distributions for the overexpressed Androgen Receptor (AR) and Forkhead Box Protein A1 (FOXA1) are broad, with mean values exceeding the threshold for direct E-P contact. Importantly, the average E-P distances decrease in the cancer cells, compared to normal cells in the AR locus, whereas in FOXA1 they are roughly constant or increase modestly. Similarly, the number of multiway contacts increases as cancer progresses across cancer cell lines in the AR locus. In contrast, the number of multiway contacts in FOXA1 is similar in normal and cancer cells. Because the 3D characteristics do not explain the enhanced gene expression in cancer cells, we developed Activity-by-Multiway-Contact (AMC) Model that integrates the multiway contacts with enhancer biochemical activity. The AMC model provides a plausible mechanism for the overexpression of both AR and FOXA1 in prostate cancer. Moreover, using Micro-C data for breast cancer, we show that the number of multiway enhancer-promoter contacts increases in four of five genes studied. When multiway contacts are combined with biochemical activity, changes in gene expression found in the experiment, positively correlate with the AMC score. The predictions of the AMC model not only account for overexpression of genes in prostate and breast cancer but also provide a basis for understanding gene expression variations in other genes as well.

genomics↗

bioRxiv2026

MitoAtlas: a Domain-Resolved Spatial Map of the HumanMitochondrial Proteome

Mitochondrial function depends on the precise sub-organellar localization and topological orientation of its proteome. However, mapping this architecture with domain-level precision across the double-membrane system has remained challenging. Here, we present MitoAtlas, a spatial systems biology resource built on SR-PL (super-resolution proximity labeling) that resolves the sub-mitochondrial architecture of 861 proteins at domain-level resolution. By deploying 42 spatially restricted APEX2 and BioID/BioID2 baits across mitochondrial compartments and cytosol in HEK293T-REx cells, we mapped 13,440 unique proximity-labeled sites. Integrating high-density spatial data with a machine-learning classification pipeline and AlphaFold structural modeling, we resolved membrane topologies for 196 transmembrane proteins and assigned sub-mitochondrial localizations to 160 previously unannotated mitochondrial proteins absent from MitoCarta3.0, including MINPP1 as an intermembrane space resident. Coupling bait-specific proximity labeling signatures with AlphaFold structure prediction and metabolite docking identified 47 protein-protein interactions, including the LETM1-Citrate Synthase complex, and 155 protein-metabolite interactions. MitoAtlas is available at mitoatlas.org as a domain-resolution spatial map of mitochondrial architecture and metabolism.

cell biology↗

bioRxiv2026

A druggable redox switch on SHP1 controls macrophage inflammation

Immunological proteins are major disease targets, yet most remain undrugged. Post-translational redox modification of cysteine residues has emerged as an important mode of immune cell regulation, particularly in macrophage cytokine responses. Here, we develop a strategy for systematic discovery and small-molecule functionalization of redox-regulated cysteines on immunological proteins. Using deep redox proteomics, we annotate 788 in vivo redox-regulated cysteines across diverse immune-relevant protein domains. We demonstrate how these sites enable cysteine-directed pharmacology through discovery of a novel cysteine activation site on the immune regulator SHP1. Targeting Cys102, we develop a highly selective covalent agonist, SCA, which binds the N-SH2 domain to relieve autoinhibition and activate SHP1. In mouse and human macrophages, SCA selectively engages SHP1 Cys102, antagonizing IRAK signaling and LPS-induced pro-inflammatory cytokine production. Together, this work identifies a druggable cysteine redox switch controlling macrophage cytokine responses and provides a compendium of redox-regulated sites for therapeutic development.

biochemistry↗

bioRxiv2026

Virus-like particle delivery enables orthogonal genome editing in vitro and in vivo

The advent of CRISPR-Cas systems has revolutionized multiple fields, including basic science, biotechnology, and medicine. Central to this versatility is the use of programmable guide RNAs (gRNAs), which enable flexible and specific gene targeting. Building on this principle, various CRISPR-associated tools have been developed, including Cas9, base editors, prime editors, and gene-regulation platforms. However, because most CRISPR modalities share a common gRNA platform, the simultaneous use of multiple tools is constrained by interactions among different gRNAs, limiting orthogonal genome editing. This study presents a method for orthogonal multiplexed gene editing by packaging distinct CRISPR effectors and their corresponding gRNAs into separate virus-like particles (VLPs), thereby virtually eliminating the gRNA crosstalk. Furthermore, we demonstrate that this VLP-based strategy enables orthogonal, multiplexed gene editing in vivo in mouse eyes and ears. This platform expands the CRISPR toolkit by enabling simultaneous, non-interfering genetic manipulations in both cultured cells and living organisms.

genetics↗

bioRxiv2026

Environmental Amino Acid Sensing Regulates the Rate of ASC Translation and NLRP3 Inflammasome Assembly

The NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome is a multiprotein signaling complex that triggers pyroptotic cell death and interleukin (IL)-1 family cytokine release during infection and cell injury. Its assembly is driven by the adaptor protein, apoptosis-associated speck-like protein containing a CARD (ASC), whose filamentation forms a supramolecular speck upon NLRP3 activation to amplify inflammasome signaling. While the NLRP3 inflammasome is well appreciated as a sensor of environmental danger and damage, little is known about how homeostatic environmental factors like dietary metabolites regulate its activity. Here, we find that environmental availability of the branched-chain amino acids (BCAAs), leucine, isoleucine, and valine, controls NLRP3 inflammasome assembly. While ASC is typically viewed as a constitutively expressed, unregulated inflammasome component, we find that Toll-like receptor 4 (TLR4) activation triggers localization of ASC mRNA to the perinuclear space. Moreover, our data demonstrate that ASC undergoes TLR4-driven translational bursting from polyribosomes during inflammasome priming. This translational engagement is dependent on BCAA availability and mechanistic target of rapamycin (mTOR) activity, which regulate the kinetics of inflammasome assembly. In contrast, the translation of NLRP3 and caspase-1 is largely insensitive to these inputs. Furthermore, we find that BCAAs regulate NLRP3 inflammasome activation in both mouse and human macrophages, in the context of bacterial infection, and during lipopolysaccharide (LPS)-induced sepsis in vivo. Altogether, this work unveils a novel inflammasome priming event governed by the amino acid environment. These findings further highlight how the activity of proteins maintained in equilibrium like ASC can be dynamically regulated through rapid changes in mRNA translation.

immunology↗

bioRxiv2025

Diverse infections transcriptionally reprogram the intestinal epithelium and epithelial-immune cell interactions

The distal small intestine plays vital roles in host physiology by regulating nutrient and fluid homeostasis. Despite being impacted in Crohns disease and a major target for a range of infections, we know relatively little about the complexity of cellular responses and cell-cell communication in the ileum during infection. Single cell and spatial transcriptomics have emerged as powerful technologies to study tissue heterogeneity in the gut, but these tools have focused on the large intestine, in part due to the accessibility of this tissue for biopsies and its importance in cancer. Here we present GutPath, an atlas of over 500,000 single cells with RNA and protein expression profiles for 91 cell states in the ileum across diverse infectious archetypes. We show that GutPath accurately captures established immune responses to infection while revealing pathogen-specific responses in enterocytes. To highlight the discovery potential of this atlas, we identify a novel enterocyte cell state present during Yersinia pseudotuberculosis infection that is spatially linked to bacterial load and tissue pathology. GutPath establishes a much-needed resource for the immunology community that will accelerate the study of the transcriptional diversity of cellular landscapes in the small intestine.

immunology↗

bioRxiv2025

Human bone marrow adipocytes drive prostate cancer bone metastasis progression via lipid-mediated induction of Angiopoietin-like 4

Bone is the main metastatic site in advanced prostate cancer (PCa) and contains bone marrow adipocytes (BMAds), which account for more than 70% of adult bone marrow. However, their role in the progression of PCa metastases remains poorly understood. Herein, we developed a physiologically relevant 3D culture model using primary human BMAds from red hematopoietic rich-areas (rBMAds) and we showed that rBMAds engage in metabolic crosstalk with PCa cells. Specifically, rBMAds release free fatty acids (FFAs) through a non-canonical lipolytic pathway and these FFAs are taken up by PCa cells inducing a transcriptional reprogramming that promotes motility. Among the responsive genes, Angiopoietin like 4 (ANGPTL4) is the most upregulated via a peroxisome proliferator-activated receptor {gamma}-dependent mechanism, and its silencing abolishes the rBMAd-driven migratory and invasive phenotype. Clinically, elevated ANGPTL4 expression is associated with poor overall survival specifically in PCa bone metastases, is maintained during metastatic progression, and is indicative of aggressive disease. Together, our findings uncover a novel adipocyte-tumor interaction and identify ANGPTL4 as a key mediator and potential therapeutic target in bone-metastatic PCa.

cancer biology↗

bioRxiv2025

Red bone marrow hosts metabolically active anucleate adipocytes that support hematopoiesis

Bone marrow adipocytes (BMAds) are a major component of the bone marrow (BM) that regulate bone turnover and hematopoiesis. In rodents, two distinct adipocyte populations exist: constitutive BMAds (cBMAds), located in areas devoid of hematopoietic cells and resistant to metabolic cues, and regulatory BMAds (rBMAds), interspersed within hematopoietic niches and responsive to energy stress. Despite their potential importance, rBMAds have remained poorly characterized due to their scarcity in rodents. Here, we used a high-yield method to isolate human rBMAds, enabling structural, proteomic, lipidomic, and functional analyses. Remarkably, human rBMAds are anucleate yet retain organelles and maintain lipid and glucose metabolism, but unlike their rodent counterparts, they lack lipolytic activity. They actively secrete factors that support hematopoiesis in vitro, implicating them as functional contributors to the BM niche. These findings suggest that human rBMAds may arise from cBMAds through terminal differentiation and define a novel adipocyte subtype critical for BM homeostasis.

cell biology↗

bioRxiv2025

Lysosome-dependent nutrient scavenging underliesstress adaptation during epithelial-to-mesenchymaltransition

Metastatic cancer cells invade tissue, overcome nutrient stress, and survive transit to distant sites. Many of the mechanisms that support these processes are incompatible with proliferation. This study defines cellular transition states in breast epithelial cells undergoing epithelial-mesenchymal transition (EMT) driven by ERK2 and TGF-{beta} signaling. EMT triggers robust endolysosomal system upregulation and metabolic adaptations that balance proliferative and invasive states. Surprisingly, invasive cells rely on scavenging via lysosomes and macropinocytosis to acquire amino acids, rather than plasma membrane transport, even in nutrient-rich conditions. Macropinocytosis increases intracellular amino acid storage, promoting survival during amino acid deprivation. This metabolic shift depends on c-MYC downregulation, an early EMT event. Reintroducing c-MYC suppresses the metabolic switch, endolysosomal induction, macropinocytosis, and the proliferation-to-migration transition. These findings reveal how cells dynamically balance proliferation and invasion, offering insights into transition states difficult to capture in models of breast cancer metastasis.

cell biology↗