Biology subjects

Kim, T.

Publications and source records attributed to Kim, T..

At least 19 recordsLinked to original sources

bioRxiv2026

Predicting conformational changes induced by biomolecular interactions

Biomolecular interactions trigger structural changes of protein complexes essential for cellular signaling, regulation, and drug action. Modern co-folding models can now generate detailed structures of multi-component complexes directly from sequence. However, recent evaluations reveal that they frequently fail to capture specific conformational states induced by a ligand or partner. Here, we show that explicitly modeling the structural transition from an unbound to a bound state, rather than generating the bound complex alone, better resolves these interaction-induced states. Our generative learning framework, K-Fold, transforms independently predicted unbound component priors into an assembled complex through an ultrafast diffusion bridge, operating entirely without multiple sequence alignments. By eliminating co-evolutionary dependencies, K-Fold accurately recovers non-coevolved interfaces in antibody-antigen and targeted-degrader complexes, as well as ligand-directed regulatory states in GPCRs and kinases. Furthermore, by coupling this diffusion bridge with diverse monomeric equilibrium ensembles, K-Fold provides a computationally tractable foundation for sampling complex multi-state landscapes without retraining. These results demonstrate that formulating molecular assembly as an explicit structural transition moves structure prediction beyond static geometries toward capturing biomolecular dynamics.

molecular biology↗

bioRxiv2026

Characterization of Resolved, Localized, and Progressing Infections at the Maternal-Fetal Interface in a Nonhuman Primate Model

Background: Inflammation and infection are responsible for the majority of early preterm births. Host innate immune responses at the maternal-fetal interface that contribute to sterile intra-amniotic inflammation (SIAI), bacterial clearance, or invasive infection remain poorly understood. Group B Streptococcus (Streptococcus agalactiae) is a leading cause of preterm birth, stillbirth, and invasive neonatal disease. Objective: To determine the host innate immune factors and transcriptional profiles that characterize the persistence versus resolution of a Group B Streptococcus infection at the maternal-fetal interface in a nonhuman primate (pigtail macaque, Macaca nemestrina) model. We hypothesized that a highly inflammatory profile, associated with interleukin-1 signaling, is linked to progression, whereas a mild or non-inflammatory signature is associated with infection resolution. Study Design: In a chronically catheterized pregnant pigtail macaque model, Group B Streptococcus was inoculated into the choriodecidual space. Infections were classified as progressing (invasive into the amniotic fluid), localized (bacteria persisting at the inoculation site), or resolved (minimal to no bacteria detected). Temporal and spatial RNA and protein profiles in the chorioamniotic membranes and decidua and in the myometrium were assessed by histopathology, multiplex protein assays, total RNA-sequencing, Nanostring nCounter, and spatial transcriptomics (digital spatial profiling). Differential gene expression was analyzed with limma using moderated t-tests and by gene set enrichment analysis. Results: A GBS infection at the maternal-fetal interface produced three infection outcomes: progressing infections that disseminated into the amniotic fluid (AF, N=5), low-titer localized infections remaining at the inoculation site (N=6), and resolved infections (N=5), compared with saline controls (N=3). Preterm labor occurred frequently regardless of infection outcome (40-80%). In resolved infections, SIAI occurred in the AF and lymphocytes infiltrated the myometrium. Total RNA-Seq of the chorioamniotic membranes demonstrated that 99 genes separated progressing from resolved infections, 98 of them were upregulated and dominated by IL-1 signaling and granulocyte recruitment (i.e., IL1B, CXCL8, TLR2, CLEC4A). Histone deacetylase 10 (HDAC10) was the only significantly downregulated gene in the progressing versus resolved contrast (p<0.05). An exploratory spatial transcriptomics experiment localized the progressing inflammatory response to the chorion. In the myometrium, zinc and RING finger 3 (ZNRF3) was significantly downregulated in localized versus control and resolved versus control and was also the only differentially expressed gene. Downregulated WNT signaling and upregulated lymphocyte-mediated immunity gene sets in the myometrium were a signature of resolved infections. Conclusion: A GBS infection at the maternal-fetal interface can result in a spectrum of progressing, localized and resolved outcomes. An early IL-1 and granulocyte response in the chorion layer of the chorioamniotic membranes marks bacterial dissemination, with loss of HDAC10 suggesting epigenetic regulation within the membranes with progressing infections. In the myometrium, ZNRF3 and downregulation of WNT signaling persisted even after infection resolution, reflecting an infectious exposure. Unexpectedly, resolved infections were associated with SIAI and a myometrial inflammatory response, offering a possible explanation for preterm labor associated with SIAI.

systems biology↗

bioRxiv2026

Evaluation of the Translational Readiness of Pluripotent Stem Cell-Derived Vascular Cell Therapy for Limb Ischemia: A Systematic Review and Meta-analysis

Background: For the first time, pluripotent stem cell (PSC)-derived endothelial cells were administered to a human for peripheral artery disease (PAD), in August 2026. This translation is based on a large preclinical literature claiming that donor cells build blood vessels in animal models of limb ischemia, but whether it supports that claim has not been assessed systematically. Methods: In this systematic review and meta-analysis, we searched MEDLINE, Web of Science, Europe PMC, and Embase to August 2026 for studies of PSC-derived vascular cells in animal models of limb ischemia with an acellular control. Limb perfusion at each experiment's primary timepoint was pooled as Hedges' g in a three-level random-effects model with cluster-robust variance. Donor-cell fate and a seven-level ordinal of vascular contribution were graded for every study. This meta-analysis was registered on PROSPERO (CRD420261465308). Results: Sixty-eight studies in 69 reports met the criteria; 51 contributed 114 comparisons at primary timepoints. Cell therapy increased perfusion (Hedges' g 2.26, 95% CI 1.72-2.79; P=4.1x10-11; I2=87.5%; 95% prediction interval -1.47 to 5.98). No prespecified moderator survived Holm-Bonferroni adjustment, including the donor-cell fate grade (P=0.30). A donor cell was resolved in a vessel wall in 35 studies, and flow through such a vessel in 7. Small-study effects were strong (Egger t=6.25, P=9x10-8) but were not reproduced by a sample-size-based test (P=0.80); trim-and-fill moved the estimate to 1.15. On the natural scale, treated limbs recovered about twice the control perfusion (ratio of means 1.96, 95% CI 1.66-2.33). Limb preservation favored cell therapy in 11 studies with countable events (risk ratio 4.88, 95% CI 2.28-10.45), but 43 of 68 studies reported no functional outcome, and only 22 comparisons from 8 studies were both randomized and blindly assessed. In a post hoc analysis that did not survive multiplicity adjustment, effects were largest in studies whose images could not resolve their incorporation claims (g 3.23) versus those that resolved incorporation (g 2.33). Conclusions: PSC-derived vascular cell therapy improved limb perfusion in animal models, consistently across every sensitivity analysis, although the magnitude is uncertain given heterogeneity and small-study effects. Benefit was not associated with the degree of donor-cell incorporation across studies and was largest where incorporation was least resolvable, a pattern more consistent with reporting and measurement bias than with a true mechanistic effect. The first rigorous, blinded test of that benefit may come not from animals but from the patients now receiving these cells.

cell biology↗

bioRxiv2026

Maternal vaccination timing shapes the composition and maturation of antibody clonotypes transferred to the newborn

Newborns rely on maternally transferred antibodies for immune protection, acquired across the placenta and through breast milk. Despite this importance, how closely the inherited antibody repertoire resembles the mothers has not been examined at the clonotypic level. We combined BCR-Seq with proteomic Ig-Seq to track SARS-CoV-2-specific antibody clonotypes across maternal blood, cord blood, and breast milk from six mRNA-immunized pregnant individuals. Vaccination earlier in gestation generated more diverse peak IgG repertoires but greater contraction before delivery, yielding fewer transferred clonotypes. Vaccination later in gestation produced more restricted peak repertoires, but more clonotypes persisted to delivery and transferred to cord. Infant cord was enriched for persistent, highly somatically mutated, and intraclonally diverse IgG clonotypes, consistent with preferential transfer of affinity-matured antibodies. In contrast, breast milk IgA repertoires were largely distinct from systemic repertoires and underwent substantial remodeling despite stable antigen titers. These findings define molecular determinants of passive immunity shaped by vaccination timing relevant to optimizing maternal immunization.

immunology↗

bioRxiv2026

BR-bodies link RNA homeostasis to stress tolerance and intracellular fitness in Brucella

Bacterial gene expression depends on the coordinated regulation of RNA synthesis, processing, and decay. The conserved RNA degradosome scaffold, Ribonuclease E (RNase E), assembles into phase-separated bacterial ribonucleoprotein bodies (BR-bodies) through its C-terminal intrinsically disordered region (IDR). This IDR also scaffolds recruitment of degradosome client proteins that carry out post-transcriptional gene regulation. Whether BR-bodies regulate virulence programs to support infection, however, is largely undefined. We show that RNase E of the intracellular pathogen Brucella ovis forms RNA-dependent condensates in vivo and phase-separates with RNA in vitro, with the IDR necessary and sufficient for BR-body assembly. Deleting the IDR (rne({Delta}IDR)) did not impair growth but sensitized Brucella to host-relevant oxidative and cell-envelope stressors. Transcriptome-wide profiling that simultaneously resolved mRNA decay and processing revealed that BR-bodies primarily accelerate mRNA turnover while stabilizing a distinct subset of transcripts. Notably, BR-bodies control the processing and levels of virB type IV secretion system (T4SS) mRNA and the turnover of its key activators, linking condensate-based RNA regulation to a core virulence pathway. Consistent with virB dysregulation, the rne({Delta}IDR) mutant was severely attenuated in mammalian macrophages. To test whether phase separation is sufficient for BR-body function, we replaced the B. ovis IDR with the highly divergent Caulobacter crescentus IDR. This chimera assembled BR-bodies but rescued fitness incompletely, fully restoring oxidative-stress resistance but not cell-envelope stress resistance or intracellular fitness. BR-bodies therefore link RNA metabolism to Brucella stress resistance and infection, and their full function requires both phase separation and additional native IDR-specific activities such as degradosome interactions. ImportanceBiomolecular condensates are non-membrane bound organelles that organize biological processes across all domains of life, but their role in bacterial infection is not well characterized. We show that Brucella, an intracellular bacterial pathogen and causative agent of the disease brucellosis, relies on biomolecular condensates called BR-bodies to control RNA stability and regulate gene expression. Without BR-bodies, Brucella is sensitized to host-relevant chemical stresses, cannot properly regulate essential infection machinery, and has severely diminished fitness within mammalian host cells. These results indicate that control of RNA levels by biomolecular condensates is required for Brucella survival within animal hosts, providing evidence that phase separation is a fundamental mechanism underlying Brucella adaptation to the hostile environment encountered during infection.

microbiology↗

bioRxiv2026

Borrelia burgdorferi infection-enhanced Ixodes scapularis serpins S12c5 and S13c5, target MASP1/3 to inhibit MBL-pathway complement activation and promote spirochete survival

Tick serine protease inhibitors (serpins) are critical for tick feeding success and pathogen transmission. We previously demonstrated that two unique serpins, S12c5 and S13c5, are tandemly duplicated on Ixodes scapularis chromosome 5 and are injected into the host at heightened levels during feeding by Borrelia burgdorferi (Bb)-infected nymphs. Here, we characterize the biochemical properties and immunomodulatory functions of these serpins and investigate their roles in promoting Bb transmission. Despite their response to Bb infection, S12c5 and S13c5 share less than 20% overall amino acid identity and differ critically at their reactive center loop (RCL) P1 site, which determines substrate specificity: S13c5 contains a basic arginine residue, while S12c5 contains a polar serine residue. Consistent with these differences, recombinant S13c5 efficiently inhibited innate immune proteases including fXa, plasmin, and trypsin IV (SI: 2.5, 1.9, and 1.0; ka: 8.56 x 103, 3.97 x 10, and 1.08 x 10 M-{superscript 1}s-{superscript 1}, respectively), and consequently delayed plasma clotting via the common pathway, whereas rS12c5 did not inhibit these proteases. However, despite molecular modeling predicting stronger interactions between S13c5 and complement proteases, both serpins inhibited MBL-pathway complement activation by targeting MASP1/3. Deletion of the RCL domain abolished this inhibitory activity, confirming both proteins function as typical inhibitory serpins. Functionally, rS13c5 enhanced Bb colonization in C3H mouse organs. Interestingly, immune response assays revealed a trade-off: S13c5, despite its broader inhibitory activity, was less immunogenic than S12c5. Collectively, these findings establish S12c5 and S13c5 as Bb transmission factors that promote spirochete survival through disruption of host innate immunity.

Molecular Biology↗

bioRxiv2026

A human developmental and adult brain atlas benchmarks dopaminergic stem cell models and cell therapy candidates

Parkinsons disease (PD) is characterized by the progressive loss of midbrain dopaminergic (mDA) neurons1. Stem cell-derived mDA neurons hold promise for disease modelling2,3 and are currently in clinical trials for cell replacement therapy4-6. However, systematic benchmarking has been limited by the lack of a unified high-resolution reference and methods that quantify incomplete or mixed lineage specification in vitro7. We establish a single-cell and spatial atlas of the human developing diencephalon-midbrain-hindbrain axis resolving 93 cell subtypes, including 39 lacking prior single-cell characterization and 4 entirely novel populations. Using this atlas as a reference, we integrate 19 hPSC-derived mDA datasets, both published2,8-25 and unpublished, to build the Human Dopaminergic Neural Atlas (HDNA) spanning 2D, 3D, and graft models, including those used in clinical trials. To classify cells and quantify lineage fidelity, we develop CapybaraBrain, a marker-driven non-negative decomposition framework that assigns each cell continuous identity scores across all 93 developmental programs, enabling systematic discrimination of discrete, transitioning, and cross-lineage hybrid states26. We uncover a pervasive landscape of off-target populations reflecting relaxed transcriptional boundaries in vitro, including a previously unrecognized TH-PITX2 midbrain neuronal population, and we validate atlas-predicted latent lineage plasticity through inducible genetic fate mapping in mouse models. We further define maturation-associated transcriptional programs by harmonizing adult mDA subtype atlases, revealing that dopaminergic identity and maturation are partially decoupled across protocols. Finally, projecting PD patient-derived tri-cultures onto the HDNA uncovers genotype- and cell-type-specific transcriptional dysregulation. Together, these integrated atlases and computational framework establish a unified standard for benchmarking differentiation fidelity, exposing off-target states, and guiding next-generation PD models and cell therapies.

Developmental Biology↗

bioRxiv2026

Mechanical Checkpoint for Cell Division in Three-Dimensional Microenvironments

Cell division within mechanically confining extracellular matrices (ECMs) is a key regulator of tissue morphogenesis and cancer progression. Although the intracellular force-generation mechanisms that drive volumetric growth and mitotic elongation are well characterized, how ECMs resist these forces remains poorly understood. Unlike linearly elastic materials, fibrillar ECMs exhibit nonlinear and viscoelastic behaviors that fundamentally alter how they oppose cell-generated stresses. Using a fiber-level computational model, we dissected the origins of ECM-mediated mechanical confinement during mitosis. We identified three distinct modes of resistance: compressive resistance at the cell poles, shear resistance from a pericellular shell, and tensile resistance at the cell equator. The relative contributions of these modes depended on fiber architecture and connectivity; however, shear resistance from the pericellular shell--pre-tensed by volumetric growth during G1--consistently dominated as the primary mechanical barrier to mitotic elongation. These findings suggest that the pericellular shell functions as a natural mechanical checkpoint on cell division within collagen-rich microenvironments. Notably, a finite element continuum model, despite being the most widely used framework for tissue mechanics, failed to reproduce these behaviors, underscoring the necessity of fiber-resolution approaches. We propose that overcoming this mechanical checkpoint is a critical step in cancer progression, enabling cells to divide within the dense stromal matrices characteristic of metastatic tumors.

biophysics↗

bioRxiv2026

Human meningiomas arise from developmental meningeal mosaicism

Meningioma is the most common primary intracranial tumour, yet its genetic origin and the temporal sequence of mutational events remain poorly defined. Here, we analysed 80 triple-matched tumour, histologically normal meninges, and blood samples from 22 patients. NF2 or TRAF7 driver mutations are detectable in phenotypically normal meninges in 81.8% of cases (95% CI: 59.7-94.8%; VAF [~]0.02%), corroborated by high-depth sequencing, single-cell cloning, and phylogenetic analysis. By distinguishing developmental mosaic mutations from postnatal tumour-private mutations, we revealed distinct mutational signatures and resolved the temporal sequence of meningioma evolution. The developmental origin was further underscored in patients with multiple meningiomas, where identical driver mutations were shared across genomically distinct tumours and meninges, and in intraventricular meningiomas (IVM), where driver mutations were detected in distant cranial dura. Reconstruction of mutational timing revealed lineage-specific trajectories, linking mosaicism to diverse disease presentations including solitary meningioma, IVM, meningiomatosis, and NF2-related schwannomatosis. Together, these findings reveal an early origin of human meningioma, in which developmental mosaicism establishes a pre-neoplastic field within the meninges, providing a developmental framework for adult tumourigenesis.

Cancer Biology↗

bioRxiv2026

Vascular Deformation Mapping Calibration with Physics-based Synthetic Data on Multi-axial Aortic Motion

Aortic stiffness is both heterogenous and anisotropic. Current non-invasive methods to estimate aortic stiffness are limited to characterizing the aortic tissue as isotropic due to the lack the techniques required to extract multi-axial strain from 3D dynamic images. Vascular deformation mapping (VDM) is a nonrigid image registration technique which has thus far been applied to map aortic growth using longitudinal imaging. In this study, we propose to use VDM to assess 3D aortic deformation by mapping diastolic and systolic images. During image registration process, penalty parameters are employed to fine-tune image alignment and penalize non-physiological deformations. These penalty parameters must be calibrated to ensure that VDM successfully reproduces multi-axial aortic motion patterns in health and disease. In this paper, we developed a calibration pipeline for these parameters using synthetic data. A rotation-free shell model was used to generate physics-based synthetic data on aortic motion incorporating patient-specific geometries, root motion, and blood pressure from a cohort of 14 subjects (healthy, Marfans syndrome and thoracic aortic aneurysm). An error metric was defined to quantify the quality of the VDM results. Furthermore, a k-means clustering technique was used to categorize the subjects into three clusters based on ascending aortic motion. Optimal penalty parameters were identified for each of the three clusters. The results indicated that patient clusters with smaller aortic root motion required larger rigidity penalty values. The calibrated parameters successively reduced errors in 3D displacement and multi-axial stretch compared to un-optimized VDM predictions, enhancing the accuracy of capturing aortic deformation from dynamic images. Among the different aortic regions, the ascending thoracic aorta exhibits the largest error reduction.

bioengineering↗

bioRxiv2026

Durotactic Migration Driven by Anisotropic Matrix Stiffening and Mechanical Feedback

Cell migration is fundamental to various biological processes, including morphogenesis, wound healing, and cancer metastasis. Durotaxis--directed migration of cells in response to spatial variations in stiffness--has been extensively studied using engineered substrates with prescribed stiffness. However, recent work has increasingly shifted toward understanding cell migration in fibrous matrices that can be actively remodeled by the actomyosin contractility, as commonly observed in tumor and epithelial cells. Despite these advances, a theoretical framework explaining how cells structurally remodel their surrounding matrix to promote their own durotaxis, and which cellular forces govern this behavior, remains elusive. To address this gap, we developed a biomechanical model in which polarized cells contract and migrate over a fibrous matrix. Using this model, we first confirmed that cells on an externally strained matrix preferentially migrate along the direction of applied strain. Then, we investigated how cells autonomously remodel the matrix to create stiffness patterns favorable for durotaxis. In the presence of intercellular adhesion, cells acted collectively to stiffen the matrix, after which a small subset of cells escaped the main population and migrated outward. This behavior is reminiscent of intravasation during cancer metastasis, where cohesive cell clusters generate local matrix remodeling that facilitates the departure of more motile subpopulations. These results illustrate how matrix stiffening driven by cell cohesion and contractility regulates durotactic behavior and provide mechanistic insight into collective invasion processes relevant to cancer metastasis.

biophysics↗

bioRxiv2026

A direct forcing immersed boundary method for biofluid simulations using a non-linear rotation free shell model on unstructured grids

Thin structures such as heart valves and aortic dissection flaps interact dynamically with blood flow in human vessels. Their flexibility and capacity for large deformations generate complex, highly transient hemodynamic patterns over the cardiac cycle. Accurately resolving these interactions remains challenging for conventional boundary-fitted fluid-structure interaction approaches. We present an immersed boundary method for simulating thin structures in incompressible flow on unstructured grids. The method couples a stabilized finite element fluid solver with a nonlinear, rotation-free shell formulation through a direct forcing immersed boundary approach. The framework supports both weak (explicit) and strong (implicit) time-coupling strategies, enabling stable simulations over a wide range of solid-to-fluid density ratios. Hydrodynamic forces acting on thin structures are computed from fluid solutions sampled on both sides of the structure, allowing accurate force reconstruction for zero-thickness shells. To our knowledge, this is the first immersed boundary formulation that couples an unstructured finite element fluid solver with a two-dimensional, rotation-free shell model to simulate interactions between thin structures and incompressible flow. Fluid-structure coupling is achieved using predefined finite element shape functions, which provide consistent projection between Eulerian and Lagrangian fields without additional interpolation procedures. The framework is validated using three-dimensional benchmark problems involving thin structures. Then, valve-like model is used to compare strong and weak coupling strategies. Finally, the method is applied to an idealized type-B aortic dissection model. The proposed approach is implemented within the open-source software CRIMSON, a finite element platform for cardiovascular simulation.

bioengineering↗

bioRxiv2026

A genome-wide CRISPR screen defines host determinants of early Brucella infection in human macrophage-like cells

Brucella spp. are widespread intracellular animal pathogens that cause brucellosis, a significant zoonosis. Despite the global impact of brucellosis on animal and human health, the host genes that support Brucella infection remain incompletely defined. To address this knowledge gap, we developed a flow cytometry-based infection assay with fluorescent Brucella and performed a genome-wide CRISPR-Cas9 loss-of-function screen in human macrophage-like cells. Disruption of >150 host genes significantly reduced intracellular B. abortus burden at 3 h post-infection. In addition to recovering known host factors, the screen revealed previously unappreciated genes linked to endosomal trafficking, cytoskeletal remodeling, and lipid homeostasis. The screen was robust, as validation within these functional categories confirmed that the small GTPase RAB14, the Src-family kinase regulator CSK, and the phospholipid flippase subunit TMEM30A support early infection by B. abortus and B. ovis without impairing general phagocytosis. Gene set enrichment analysis further revealed positive regulators of mTORC1 signaling as key host factors; this result was validated through targeted disruption of LAMTOR2 and AKT1, and pharmacologic inhibition of AKT1. Thus, the AKT-Ragulator-mTORC1 signaling axis contributes to the establishment of a permissive intracellular niche during early Brucella infection. Finally, to assess whether these host requirements extend beyond Brucella, we examined infection by the unrelated intracellular pathogen Mycobacterium abscessus. CSK, AKT1, and LAMTOR2 were required for efficient M. abscessus infection, whereas RAB14 was dispensable. Together, these results define host genes that support early Brucella infection and distinguish shared versus pathogen-specific host dependencies exploited by intracellular bacteria.

microbiology↗

bioRxiv2026

Transgene Expression Kinetics and Replication Potential of Recombinant Adenovirus Serotype 4 in a Mouse Model and its Use as a Herpes Simplex Virus Vaccine

Human adenovirus serotype 4 (Ad4) is used as a replication-competent oral vaccine that safely and effectively prevents Ad4 respiratory illness in US military personnel. Recombinant Ad4 vaccine candidates elicit mucosal and systemic immune responses against respiratory viruses in hamsters, nonhuman primates, and humans. Although evaluation of Ad4 vaccine candidates in mice would be extremely useful given the large number of immunologic tools available, this has been limited by concerns about a lack of viral replication in these animals. Here we generated recombinant Ad4 vectors that express either luciferase (Ad4-Luc) or herpes simplex virus type 2 (HSV-2) glycoprotein D (Ad4-gD2) to identify transgene expression kinetics, the presence of Ad4 vector replication, and HSV-2 immune responses and protection against HSV-2 infection. Local luciferase activity was observed from 7 hours to 20 days after intranasal inoculation of BALB/c and humanized mice. Subsequent inoculations with Ad4-Luc showed reduced luciferase expression in BALB/c mice, but robust expression in humanized mice, suggesting an immune response to the vector in wild-type mice. Ad4 DNA, but not luciferase activity, was reduced in the lungs of BALB/c mice treated with cidofovir before inoculation with Ad4, implying that Ad4 replicated, albeit at a low level, in the lungs. Intranasal vaccination of mice with Ad4-gD2 resulted in HSV-2 neutralizing antibody in the serum, and after HSV-2 intravaginal challenge reduced disease scores, increased survival, and reduced shedding. Overall, the BALB/c mouse model is semi-permissive to Ad4 mucosal infection, but transgene expression is sufficient for the study of Ad4-based vaccine candidates. ImportanceMucosal surfaces serve as the primary site of infection and shedding for many viral pathogens. Immune responses at mucosal sites provide protection, but few mucosal vaccines are licensed. The oral replication-competent adenovirus serotype 4 (Ad4) vaccine is used to prevent respiratory illness in military recruits, has an extraordinary record of safety and efficacy and has been tested as a recombinant platform for other viruses. Further development of this vaccine platform has been partially hindered by the perceived inability to evaluate vaccine candidates in mice. Here we characterize recombinant Ad4 transgene expression kinetics and viral replication after inoculation at various sites and show protection against herpes simplex virus type 2 (HSV-2) genital disease in mice after intranasal vaccination. We show that Ad4 can induce protective efficacy, even in a semi-permissive mouse model, suggesting this is a promising vector for HSV-2 and potentially other viral pathogens.

immunology↗

bioRxiv2026

Cerebellar induced VTA plasticity underlies chronic stress-induced depression-like behaviors

Synaptic regulation is a key mechanism underlying neural circuit homeostasis and plasticity. We previously showed that projections from the deep cerebellar nuclei (DCN) to the ventral tegmental area (VTAp-DCN neurons) contribute to depression-like behaviors following chronic restraint stress (RS), but the mechanisms by which these outputs induce downstream long-term functional changes remain unknown. Here, we show that chronic RS induces an activity-dependent reduction in vesicular glutamate transporter 2 (VGLUT2) expression in VTAp-DCN axons, resulting in decreased miniature excitatory postsynaptic current frequency in dorsolateral VTA dopaminergic neurons and reduced phasic dopamine release in the nucleus accumbens. Notably, VGLUT2 reduction temporally coincides with the emergence of depression-like behaviors. Consistent with this, targeted VGLUT2 overexpression in VTAp-DCN neurons rescues synaptic transmission and dopamine signaling and prevents depression-like behaviors following chronic RS. These findings reveal a stress-induced plasticity mechanism in the cerebellar-VTA circuit linking chronic stress to depression-like behaviors.

neuroscience↗

bioRxiv2026

Functional connectivity during drawing after upper extremity peripheral nerve surgery: enhanced connectivity between motor and visuomotor-parietal regions

ImportanceRecovery after upper extremity peripheral nerve injury (PNI) surgery depends on changes in cortical neural patterns that support sensorimotor control. Task-based functional connectivity (FC) can characterize these changes, yet few studies have explored FC during ecologically fine motor valid tasks after PNI. ObjectiveTo investigate task-based FC with the left primary motor cortex (M1) during right hand drawing in individuals following right hand PNI surgery. ParticipantsForty-four right-handed adults, including 12 patients post PNI surgery (n = 8 with nerve repair, n = 4 with nerve transfer) and 32 healthy controls. MethodsAll participants underwent fMRI while performing a RH visuomotor precision drawing task. Seed-based connectivity analysis was performed to characterize the pattern of FC between left M1 and all voxels in the brain. We hypothesized that left M1 FC would differ between patients and controls, between Repair and Transfer groups, and covary with time since surgery. ResultsPatients (vs. controls) showed greater FC between left M1 and right visual and premotor cortices. Nerve transfer (vs. repair) showed greater FC between left M1 and right inferior parietal areas. Time since surgery was not linearly related to FC, though exploratory analyses suggested a negative association between log-time and FC between left M1 and right inferior parietal lobule. ConclusionAfter PNI surgery, visuomotor precision drawing involved distinct and behaviorally relevant neural patterns, which varied by task demand and potentially by surgical group despite clinical heterogeneity. Inferior parietal cortex may be especially engaged in early months after surgery (i.e. log-time). To improve recovery of upper limb function after PNI, clinical recommendations include incorporating early function-specific dexterous training, tailoring rehabilitation across surgical and recovery stages, and using multidimensional assessments of hand function.

neuroscience↗

bioRxiv2026

DNAM-1 immunoreceptor integrates innate and adaptive immune programs to drive intestinal inflammation

Innate and adaptive immune responses play critical roles in the pathogenesis of inflammatory bowel disease (IBD), yet the molecular pathways integrating these responses remain elusive. Here, we identify DNAM-1 immunoreceptor as a central driver of colitis through distinct, cell type-specific mechanisms. Transcriptomic analyses of human and murine group 3 innate lymphoid cells (ILC3s) revealed DNAM-1 as a conserved IL-23-responsive surface molecule associated with inflammatory cytokine production. In an innate immune-driven anti-CD40 monoclonal antibody (mAb)-induced colitis model, DNAM-1 expressed on ILC3s promoted intestinal inflammation by enhancing IL-22 and GM-CSF production via the integration of the Akt-mTORC1-HIF-1 signaling pathway. Genetic ablation or antibody-mediated blockade of DNAM-1 attenuated inflammatory cytokine production and disease severity. Paradoxically, in T cell-dependent colitis, DNAM-1 expression on dendritic cells, but not on ILC3s or CD4 T cells, exacerbated disease by promoting dendritic cell activation and pathogenic Th1 and Th17 differentiation. Notably, therapeutic blockade of DNAM-1 ameliorated disease in both colitis models and exerted complementary effects when combined with anti-TNF therapy, accompanied by modulation of immune activation programs distinct from those regulated by TNF inhibition. Collectively, these findings establish DNAM-1 as a pivotal regulator of intestinal inflammation bridging innate and adaptive immunity and identify DNAM-1 blockade as a next-generation therapeutic strategy for IBD. Highlight{blacktriangleright} DNAM-1 is an IL-23-responsive receptor conserved in human and mouse ILC3s. {blacktriangleright}DNAM-1 on ILC3s drives innate colitis via Akt-mTORC1-HIF-1 signaling. {blacktriangleright}DNAM-1 on DCs promotes T cell-dependent colitis by inducing Th1/Th17 cells. {blacktriangleright}DNAM-1 blockade targets immune pathways distinct from TNF inhibition. {blacktriangleright}Combined DNAM-1 and TNF blockade shows additive therapeutic efficacy in colitis.

immunology↗

bioRxiv2026

Ca2+ influx through ER-plasma membrane contacts is required for brown fat thermogenesis and metabolic health

Brown adipose tissue (BAT) exhibits exceptional metabolic plasticity, rapidly increasing energy expenditure to sustain thermogenesis during cold exposure. This high metabolic activity imposes substantial demands on cellular systems, requiring robust adaptive mechanisms to maintain homeostasis and prevent cellular stress. Yet, the pathways that support and coordinate these adaptive responses in brown adipocytes remain incompletely understood. Here, we identify a cold-induced adaptive program in BAT characterized by the formation of endoplasmic reticulum-plasma membrane (ER-PM) contact sites and the activation of store-operated calcium entry (SOCE), which is essential for maintaining brown adipocyte health during thermogenic activation. Cold exposure enhances ER-PM contacts and upregulates the expression of STIM and Orai proteins, key mediators of SOCE. Loss of STIM in brown adipocytes disrupts intracellular Ca{superscript 2} homeostasis and induces aberrant aggregation of ER membranes. STIM deficiency also impairs cold-induced mitochondrial fission resulting in hyperfused mitochondria with reduced oxidative capacity, independently of UCP1 abundance. Importantly, mice lacking STIM in BAT exhibit impaired lipid oxidation, are cold intolerant and develop exacerbated peripheral insulin resistance when challenged with a high-fat diet. Together, these findings identify ER-PM remodeling and STIM-mediated SOCE as a central regulator that links organelle architecture to brown adipocyte function and contributes to whole-body metabolic homeostasis.

cell biology↗