Biology subjects

Han, L.

Publications and source records attributed to Han, L..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

Coronavirus envelope protein drives iron sensing disorder by hijacking the TAp73-FDXR axis

Iron overload is increasingly recognized as a critical contributor to coronavirus pathogenesis1, yet the underlying induction mechanisms remain unclear. Here, we uncover a fundamental pathway by which coronavirus drives IRP1 RNA-binding activity to induce iron accumulation2 via targeting the TAp73-FDXR axis. Specifically, coronavirus infection represses transcription of FDXR (encoding the key rate-limiting enzyme in host iron-sulfur cluster synthesis3), thereby impairing host iron-sulfur cluster generation to trigger the functional conversion of the cytosolic aconitase 1 (ACO1) into iron-regulatory protein 1 (IRP1)4, ultimately leading to the hosts persistently false perception of iron deficiency. We identify TAp73 as the primary transcription factor governing FDXR expression, and demonstrate that the coronavirus envelope protein (CoV-E) orchestrates TAp73 nuclear export. Subsequently, CoV-E binds TAp73 through a critical valine residue within its C-terminal PBM domain, inducing the K48-linked ubiquitination and proteasomal degradation of TAp73. Furthermore, we developed a CoV-E-targeting molecule, DPTP-FC, which blocks CoV-E-TAp73 interaction via forming steric hindrance and effectively alleviates iron accumulation and tissue damage caused by PEDV, PDCoV, and SARS-CoV-2 infection. Our study reveals the central role of the TAp73-FDXR axis in CoV-induced iron accumulation, highlighting CoV-E as an attractive antiviral target and DPTP-FC as a promising therapeutic candidate.

microbiology↗

bioRxiv2026

Mechano-activation of synovial fibroblasts and macrophages during OA progression in the dynamically stiffening synovial microenvironment

ObjectiveDuring osteoarthritis (OA) progression, the synovial membrane undergoes profound structural and compositional remodeling and fibrosis. We sought to elucidate how evolving synovial microenvironmental mechanics during fibrotic remodeling influence cell behavior and drive the progression of synovial pathology. MethodsSkeletally-mature male C57BL/6J mice were subjected to destabilization of the medial meniscus (DMM). To control for surgical confounders, both sham-operated and unoperated mice were included, with evaluation at 4- and 8-weeks. Synovial micromechanics were quantified via atomic force microscopy (AFM). Single-cell RNA sequencing (scRNA-seq), RNA fluorescence in situ hybridization (FISH), and flow cytometry were employed to investigate cellular heterogeneity, spatial organization, and crosstalk within fibrotic and non-fibrotic synovial niches. ResultsProgressive fibrotic remodeling and marked matrix stiffening were observed in DMM-operated synovium but absent in sham- and un-operated controls. While both sham and DMM joints mounted an acute stromal and immune response to surgery, these changes resolved over time in sham conditions but persisted in DMM synovium. During disease progression, distinct functional subsets of synovial fibroblasts and immune cells emerged, with mechanosignalling pathways and distinct immune cell-fibroblast crosstalk robustly activated within DMM-induced fibrotic microenvironments. ConclusionThis study demonstrates the complex cellular dynamics and crosstalk that differentiate the evolution of the pathological synovial response in the fibrotic DMM condition relative to surgical sham controls. Our findings highlight mechanotransduction as a central mechanism driving OA synovial pathogenesis and underscore the utility of the DMM model as a platform to dissect the molecular underpinnings of synovial fibrosis.

bioengineering↗

bioRxiv2026

A dynamic RNA hub facilitates activation induced cytidine deaminase recruitment to the immunoglobulin heavy chain locus

Activation-induced cytidine deaminase (AID) converts cytosines to uracils within the actively transcribed switch regions to initiate DNA repair and formation of DNA double strand breaks required for immunoglobulin class switch recombination (CSR). How AID specifically targets switch regions remains a key unanswered question. Using a multimodal live cell single molecule imaging approach, we demonstrate that intronic switch regions promote robust transcription by enhancing polymerase loading and persistent transcriptional bursts, resulting in the formation of a dynamic RNA hub consisting of numerous nascent switch transcripts simultaneously tethered to the IgH locus. We further demonstrate that AID interacts with switch region RNA in vivo, and that this interaction is required for recruitment of AID to the IgH locus. Together, our findings show that the RNA hub formed by nascent switch region transcripts may be part of a "class switch recombination center" and drives the specific recruitment of AID to the IgH locus to initiate CSR.

cell biology↗

bioRxiv2026

Site-specific integration of transposon via engineered piggyBac transposase

The precise integration of large DNA fragments into the human genome holds significant therapeutic potential. Here, we demonstrate that combining engineered piggyBac (PB) transposase with CRISPR/Cas9 enables targeted integration of PB transposons into specified genomic loci. Our engineered PB transposase (PBase) retains high excision activity while substantially reducing endogenous integration activity. In the developed Cas9-PBase fusion system, PBase excises the transposon to generate linear DNA fragments, while Cas9 introduces site-specific double-strand breaks (DSBs), facilitating insertion of the excised fragment at the target locus. The optimized tool achieves 6.1-7.3 kb transposon integration at multiple genomic sites with 10-15% efficiency, demonstrating 60-80% targeted integration specificity. As a proof of concept, we inserted a 7.1 kb transposon encoding three genes into the {beta}2M locus of human induced pluripotent stem cells (iPSCs), conferring protection against allogeneic natural killer (NK) cell-mediated cytotoxicity in derived iNK cells. These results establish Cas9-PBase as a precise and programmable platform for large DNA sequence insertion with potential clinical applications.

genetics↗

bioRxiv2026

Systematic fMRI signal differences across cohorts alter lifespan connectome trajectories

Large-scale lifespan neuroimaging studies increasingly integrate data across distinct cohorts to characterize trajectories of brain development and aging. However, systematic differences in acquisition protocols and hardware across cohorts can alter signal characteristics in ways that bias downstream analyses. Here, we examine three cohorts from the Human Connectome Project (HCP), spanning development (HCP-D), young adulthood (HCP-YA) and aging (HCP-A), to illustrate this issue and evaluate existing strategies to mitigate it. HCP has set standards for open, deeply phenotyped, high-resolution human neuroimaging, which are frequently used as high-quality reference datasets in tool validation, replication studies, and cross-cohort meta-analyses. However, neuroimaging acquisitions have differed across HCP cohorts because of changes in scanner hardware and acquisition sequences across study phases. Because of HCPs widespread usage, even modest protocol differences between cohorts-and their downstream effects-can have outsized impacts on the field of neuroscience research. Our analysis reveals that the HCP-YA cohort exhibits systematically weaker temporal signal-to-noise ratio (tSNR) relative to HCP-D/A. These signal quality discrepancies propagate to downstream analyses, leading to differences in overall resting-state functional correlations and whole-brain and node-level measures of resting-state network organization (e.g., system segregation, modularity, participation coefficient). Consistent with protocol-driven signal differences, resting-state network measures derived from HCP-YA depart from expected lifespan trajectories, as confirmed by examination of two other lifespan datasets. Harmonization approaches accounting for protocol and scanner-model differences substantially lessen these artifactual differences in brain network measures. Our findings underscore that signal differences do not merely introduce noise, but can qualitatively alter estimated lifespan trajectories of functional network organization, including partially inverting expected lifespan patterns. Without appropriate harmonization, analyses that combine HCP cohorts can therefore result in biologically misleading inferences about brain development and aging. We demonstrate how small acquisition differences bias resting-state-derived network metrics, and how these effects can be mitigated. This work advances best practices for valid inference in multi-cohort lifespan neuroscience research.

neuroscience↗

bioRxiv2025

IndivSTATIS: A multivariate approach to analyze brain network configurations with individualized parcellation

A critical step in the analysis of large-scale functional brain networks in neuroimaging is parcellation, which defines the nodes of a brain network. Group or atlas-based parcellation schemes use a shared common space, ensuring that each individual has the same number of brain parcels, which facilitates standard analytic approaches. However, studies reveal individual differences in the boundaries of brain areas. Extracting signals using atlas-based schemes can result in varying levels of blurring of signals across homogeneous areas within a specific individuals brain. Individualized parcellation schemes can be obtained when sufficient data are available; however, these approaches introduce a significant analytical challenge: the number of parcels and networks differ across individuals. Here, we introduce IndivSTATIS, a new multivariate method based on the STATIS framework, designed to integrate individualized parcellation schemes while maintaining comparability across participants in a shared component space. The resulting network/node component scores can be used to predict individual differences measures (e.g., age, behavior). By allowing individualized parcellations to be compared within a common component space, IndivSTATIS provides a solution for incorporating individual network variability into larger studies, with potential to improve the sensitivity and interpretability of functional brain markers across both basic neuroscience and clinical applications.

bioinformatics↗

bioRxiv2025

Deletion of the polarity protein Llgl1 impairs cardiomyocyte intercalated disc integrity and permits cell cycle activity into adulthood

BackgroundCell polarity orchestrates spatial organization within tissues influencing cellular processes such as adhesion, proliferation, and differentiation. While the role of polarity proteins is well-established in epithelial systems, their function in mammalian cardiomyocytes remains poorly understood. Global deletion of the polarity protein, Llgl1, has been shown to impair cardiac trabeculation and disrupt cardiomyocyte junctions in zebrafish. Here, we investigate the role of Llgl1 in mammalian cardiomyocyte function. MethodsWe developed a mouse model whereby Llgl1 is deleted from cardiomyocytes (Llgl1CKO) and assessed cardiomyocyte intercalated disc integrity, cell cycle activity, cardiac function, and electrophysiology. We subjected mice to myocardial infarction to test if permanent Llgl1 deletion, or inducible Llgl1 deletion in adult cardiomyocytes (Llgl1TamCKO), influences the cardiac injury response. We performed RNAsequencing and in vitro knockdown and viral overexpression experiments to delineate the mechanisms by which Llgl1 influences cardiomyocyte phenotypes. ResultsLlgl1CKO mice display disrupted intercalated disc integrity indicated by reduced protein density at cardiomyocyte junctions, associated with electrical conduction defects that manifest as ventricular arrhythmia. Furthermore, Llgl1CKO mice display enhanced cardiomyocyte cell cycle activity extending into adulthood. Llgl1CKO mice and mice whereby Llgl1 is deleted by tamoxifen induction in adulthood both show improved cardiac function and reduced scar size following myocardial infarction. Mechanistically, Llgl1 alters mechanotransduction and cell cycle pathways in the heart, and regulates proper localization of N-cadherin to the cell membrane, a process that is optimized by phosphorylation of five serine residues at the C-terminus of Llgl1. ConclusionsWe elucidate a key role for Llgl1 in establishing proper intercalated disc integrity in mammalian cardiomyocytes. Disruption of intercalated discs and mechanotransduction via Llgl1 knockout is associated with cardiac arrhythmia, elevated cell cycle activity and resistance to cardiac injury. These studies shed light on mechanisms underlying intercalated disc establishment and connect disrupted cardiomyocyte junctional integrity with enhanced cell cycle activity.

cell biology↗

bioRxiv2025

O-GlcNAc Transferase Regulates GABAergic Synapse Organization and Receptor Composition.

Neural circuits must integrate metabolic information to maintain stable activity and appropriate behavioral responses. While metabolic regulation of excitatory synapses has been well studied, far less is known about how inhibitory synapses respond to changes in energy state. In this study, we show that O-GlcNAc transferase (OGT), a dynamic sensor of cellular nutrient flux, localizes to postsynaptic sites of the inhibitory synapses where it modulates synapse morphology and receptor composition. OGT over-expression reduced the size and intensity of vGAT and gephyrin puncta, whereas conditional OGT deletion produced a converse enlargement and redistribution of inhibitory scaffolds and vesicular proteins. Furthermore, OGT deletion accelerated inhibitory postsynaptic current decay kinetics and induced subunit-specific shifts in GABAA receptor surface expression: {beta}3 subunits decreased, whereas {gamma}2 subunit total and surface levels increased. Together, these findings identify OGT as a metabolic regulator that modulates inhibitory synapse structure and signaling, providing a mechanistic link between energy state and GABAergic circuit function in health and disease.

neuroscience↗

bioRxiv2025

Bruch's Membrane Contributes to the Structural Integrity of the Human Eye

PurposeTo investigate the contribution of the Bruchs membrane and sclera tissues to the overall structural integrity of the ocular wall. MethodsTwenty-three human globes were subjected to biomechanical testing. A piece of sclera measuring 5 x 5 mm was carefully removed at the nasal region, 2 mm away from the optic nerve head. The intraocular pressure was increased at approximately 1 mmHg/s until Bruchs membrane-uvea-retina-tissue layer (BMUR) ruptured. Next, strips of sclera and Bruchs membrane-choriocapillaris (BMC) complex were isolated from the superior fundus region. Uniaxial tension tests were performed at a strain rate of 0.01/s and sampling rate of 15 Hz. The tangent moduli of the BMC and sclera at 0.01, 0.02 and 0.03 strains were compared. ResultsThe rupture pressure of the BMUR was 98.1 {+/-} 21.4 mmHg. The tangent moduli of the BMC at 0.01, 0.02 and 0.03 strains were 2.96 {+/-} 1.44 MPa, 7.68 {+/-} 1.78 MPa and 9.43 {+/-} 2.11 MPa, respectively, and the tangent moduli of the sclera at 0.01, 0.02 and 0.03 strains were 1.09 {+/-} 0.80 MPa, 2.72 {+/-} 1.67 MPa and 5.69 {+/-} 3.27 MPa, respectively. ConclusionThe BMUR was able to sustain relatively high IOP before rupturing. The uniaxial tensile tests showed that the BMC tangent moduli were about 3 times of those of the sclera at strains of 0.01 and 0.02. Although the sclera is approximately 47 times thicker, the BMC is still likely to make a significant contribution (3.51% to 7.42% at strain <0.03) to the overall structural strength of the ocular wall.

bioengineering↗

bioRxiv2025

Decorin promotes nascent proteoglycan retention in cartilage matrix by strengthening collagen II-aggrecan integration

Cartilage extracellular matrix (ECM), a hydrated collagen II-aggrecan composite, undergoes dynamic turnover during both normal homeostasis and disease-associated remodeling. This study elucidates a crucial role for decorin in promoting the retention and stability of nascent aggrecan within this matrix. By applying bio-orthogonal click-labeling, we demonstrate that loss of decorin accelerates the release of nascent aggrecan under both physiological and inflammatory conditions, without affecting its preferred localization to the pericellular matrix. Conversely, supplementation with exogenous decorin mitigates inflammation-induced loss of nascent aggrecan, supporting its potential as a therapeutic target. At the molecular level, decorin exhibits strong binding affinity for aggrecan, and enhances aggrecan-aggrecan and aggrecan-collagen II interactions, reinforcing its direct role in integrating cartilage matrix constituents. Also, by binding to collagen II, decorin stiffens the collagen II fibril network, thereby strengthening the confinement effect that limits the diffusive loss of entrapped aggrecan. Notably, decorin does not alter chondrocyte transcriptomic profiles in vivo, emphasizing its primary role in maintaining matrix integrity through biophysical mechanisms rather than cell signaling. Together, these findings provide a mechanistic foundation for developing decorin-based biomaterials or gene therapies aimed at preserving or regenerating the cartilage matrix for improved outcomes in osteoarthritis.

biophysics↗

bioRxiv2025

NEUROG3 Is Sufficient to Drive Neuroendocrine Differentiation in Prostate Cancer Cells.

Treatment-emergent neuroendocrine prostate cancer (t-NEPC) arises following androgen deprivation therapy, leading to androgen-independent growth. Although multiple factors have been shown to be necessary for neuroendocrine differentiation (NED), their sufficiency has not been demonstrated. The prostate and colorectum share a common hindgut origin, and prostate neuroendocrine cell markers overlap with colorectal enteroendocrine cell (EEC) markers. Analysis of patient datasets revealed NEUROG3 amplification in both castration-resistant and neuroendocrine prostate cancers, correlating with poor survival. Because Neurogenin-3 (NEUROG3) is necessary and sufficient for EEC differentiation in the colorectum, we hypothesized that it could similarly drive NED in prostate cancer cells. A transient pulse of NEUROG3 repressed luminal identity and activated neuroendocrine programs, producing neuroendocrine cells within seven days. In summary, our findings identify NEUROG3 as a potential mediator of prostate cancer progression and establish a rapid in vitro model in which its transient activation is sufficient to initiate neuroendocrine differentiation.

cancer biology↗

bioRxiv2025

A Population-scale Single-cell Spatial Transcriptomic Atlas of the Human Cortex

The genetic and spatial determinants of cell type diversity in human cerebral cortices remain poorly defined. Here, we present a population-level single-cell spatial transcriptomic atlas of human cortices from 71 donors across the lifespan. We identified 906 layer-specific genes showing conserved and divergent laminar expression patterns between humans and other species. Spatial analysis revealed neuronal vulnerability and glial activation during aging, together with a decline in the proportion of superficial SST neurons and their interactions with other cells. Disease-associated genes exhibited high cell-type and layer-specific expression, implicating the pathogenic role of spatially specific gene expression. Spatial cis-eQTL analysis identified regulatory variants linked to genes related to diseases like Tourette syndrome. Cross-species comparison demonstrated glial expansion in the human cortex, accompanied by enhanced neuron-glia communication via the neuregulin signaling. Together, we provide a comprehensive single-cell atlas of the human cortex that is essential for understanding aging, evolution, and disease pathogenesis.

neuroscience↗

bioRxiv2025

Role of tankyrase scaffolding in the β-catenin destruction complex and WNT signaling

Aberrant WNT/{beta}-catenin signaling drives tumorigenesis and metastasis in cancer. Although enzymatic inhibitors of tankyrase (TNKS) effectively block AXIN degradation and stabilize the {beta}-catenin destruction complex (DC), they have demonstrated limited efficacy in various cancer models. Here we demonstrate that, unexpectedly, the induction of AXIN puncta represents a major barrier to achieving therapeutic efficacy. Mechanistically, catalytic inhibition of TNKS prevents TNKS turnover and drives its accumulation in the DC, wherein the scaffolding function of TNKS induces AXIN puncta formation, rigidifies the DC, and impedes {beta}-catenin turnover. Chemically induced degradation of TNKS overcomes this limitation by stabilizing AXIN without puncta formation, providing a deeper suppression of the WNT/{beta}-catenin pathway activity and the proliferation of colorectal cancer cells harboring dysfunctional APC mutations. Collectively, these findings provide an explanation for the unsatisfactory outcomes of drugging the WNT/{beta}-catenin signaling pathway by TNKS inhibitors and highlight TNKS degradation as a promising approach to treat WNT/{beta}-catenin-driven cancers.

cancer biology↗

bioRxiv2025

Mapping Human Survivability at Extreme Wet-Bulb Temperatures 32-35°C

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=146 SRC="FIGDIR/small/677706v1_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@70b72dorg.highwire.dtl.DTLVardef@9fbfeorg.highwire.dtl.DTLVardef@17f05a5org.highwire.dtl.DTLVardef@4b977b_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG HighlightsWe report the first empirical human survival map across wet-bulb temperatures of 32-35{degrees}C. HydraAon and behavior cooling extend survival beyond the 6-hour theoreAcal limit. Females consistently tolerate extreme humid heat longer than males in controlled trials. Our findings refine heat-health models and inform equitable climate adaptaAon strategies. In briefExtreme humid heat, measured by wet-bulb temperature (Tw), is nearing the threshold of human survivability under climate change. In controlled trials with 36 adults, we provide the first empirical survival map across Tw=32-35{degrees}C, showing that hydration and behavioral adaptations extend endurance beyond the theoretical 6-hour limit, with women tolerating heat longer than men. These findings refine human heat tolerance thresholds and inform equitable climate adaptation and heat-health strategies. SCIENCE FOR SOCIETYAs climate change accelerates, extreme humid heat events--measured by wet-bulb temperature (Tw)--pose a growing threat to human survival. This study provides the first empirical benchmarks of human survivability across Tw=32-35{degrees}C, clarifying how long healthy adults can endure these conditions under full hydration and minimal activity. We find that while the theoretical 6-hour survival limit at Tw=35{degrees}C is partly supported, both men and women can withstand these conditions for 7-8 hours when hydrated, with women demonstrating greater tolerance. At Tw=34{degrees}C, projected heatstroke onset extends beyond 12-16 hours, while conditions at Tw=32-33{degrees}C remain largely compensable for more than 30 hours. These results challenge prevailing heat tolerance models, underscore the role of behavioral and physiological adaptations, and provide critical data for designing equitable heat-health policies and early warning systems that account for sex differences and vulnerable populations. Climate change-driven extreme heat events increasingly threaten human health. Here, we provide the first empirical map of human survivability across four extreme wet-bulb temperatures (Tw=32- 35{degrees}C) in controlled trials. Thirty-six healthy young adults (20 males, 16 females) were exposed to twelve dry-bulb temperature and relative humidity combinations in shaded indoor settings under full hydration. Participants remained seated and performed light office tasks until core temperature (Tcore) reached 39{degrees}C or exposures lasted 8 hours. Clinical heatstroke threshold (Tcore=40.5{degrees}C) were projected by extrapolating from our direct Tcore measurements during controlled exposure trials. At Tw=35{degrees}C, males and females reached heatstroke in approximately 7.1-7.7 and 8.3-8.6 hours, respectively--partly supporting the 6-hour theoretical limit while revealing enhanced short-duration resilience when hydrated. At Tw=34{degrees}C, projected times are 12.5-12.9 hours (males) and 15.5-16.2 hours (females). Conditions at Tw=32{degrees}C and 33{degrees}C remained (quasi-)compensable, with projected heatstroke onset after 33-35 hours. Females consistently tolerated heat stress longer than males. These physiological benchmarks are critical for improving heat-health models and embedding sex-specific vulnerability in equitable adaptation strategies amid accelerating global warming.

physiology↗

bioRxiv2025

Integrative Multi-omics Deciphers the Assembly Rules of the Human Skin Microbiome and Highlights Lipid-Mediated Niche Partitioning

Understanding the ecological assembly mechanisms of microbial communities is critical for deciphering host-microbiota interactions, however, such processes remain poorly characterized in cutaneous ecosystems. By profiling a non-industrialized rural cohort, this study deciphers the assembly rules governing the human skin microbiome and reveals an evolutionarily conserved core microbiota structured by niche selection. Microbial -diversity peaks when C. acnes and M. osloensis reach equilibrium, supporting the Intermediate Disturbance Hypothesis. Host lipids, rather than aqueous compounds, emerged as the primary drivers of microbial niche partitioning. We further demonstrate that C. acnes and Malassezia restricta function as core metabolic engineers, expressing diverse lipases that hydrolyze triglycerides (TGs) into free fatty acids (FFAs), thereby maintaining skin surface lipid homeostasis ([~]85% TG + FFA). Notably, virtually all skin surface FFAs are microbially derived and function as "public goods", enabling cross-feeding that supports auxotrophic members and fosters mutualistic interactions. Importantly, in vitro coculture experiments confirm that C. acnes establish a survival niche for M. restricta. Our findings support a paradigm shift from a taxonomic to a functional understanding of the skin ecosystem, underscoring microbial lipid metabolism as a central orchestrator of ecological stability and a promising target for therapeutic intervention. O_FIG O_LINKSMALLFIG WIDTH=182 HEIGHT=200 SRC="FIGDIR/small/674196v1_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@1274fa5org.highwire.dtl.DTLVardef@1ddfa07org.highwire.dtl.DTLVardef@49f917org.highwire.dtl.DTLVardef@807a14_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGRAPHIC ABSTRACTC_FLOATNO C_FIG

microbiology↗

bioRxiv2025

Segmented poly(A) tails with microRNA target sites confer tissue-specific regulation for mRNA therapeutics

Targeted delivery and controlled expression of mRNA-LNPs are critical for the development of safe and effective mRNA medicines. However, efficient post-delivery regulation of mRNA-LNP expression remains challenging. In this study, we engineered segmented poly(A) tail variants that function as gene-specific regulatory elements for synthetic mRNAs. Specifically, we inserted microRNA target sites (MTS) for miR-122 or miR-142 at various positions of the poly(A) tail of synthesized luciferase reporter mRNA. These modifications significantly reduced luciferase expression in non-target tissues in vitro and in vivo, demonstrating position-dependent selective expression control. Furthermore, by incorporating triple-MTS sequences for miR-142, miR-126, and miR-148a in all possible combinations at the 5 end of the poly(A) tail, we identified triple-MTS arrangements that simultaneously decrease luciferase activities in three non-target hepatic cell types, while preserving robust expression in hepatocytes. The results highlighted the importance of MTS insertion order for optimal mRNA silencing. These triple-MTS modules significantly expanded the utility of single miRNA-responsive elements, enabling cell-type selective regulation for mRNA therapeutics. By complementing tissue-tropic delivery LNPs, our mRNA cargo regulatory elements have the potential to improve tissue and cell type selectivity as a novel platform for post-delivery regulation of mRNA-LNP.

bioengineering↗

bioRxiv2025

O-GlcNAcase promotes dendritic spine morphogenesis while downregulating their GluA2-containing AMPA receptors

Dendritic spines are essential for synaptic transmission, neural circuit organization, and cognitive function. Their morphology and density influence synaptic plasticity, learning, and memory. Many proteins in dendritic spines are modified with O-GlcNAc, a monosaccharide that can be attached and removed from serines and threonines. O-GlcNAc has been implicated in multiple brain disorders, yet the role of O-GlcNAcase (OGA), the enzyme that removes O-GlcNAc modification from proteins, in dendritic spine regulation remains unclear. This study examines the role of OGA in spine and synapse morphogenesis. Immunohistochemical and biochemical analyses reveal OGA present in dendritic spines. Functional assays show that OGA promotes spine maturation, increases spine density, and alters synapse size. Additionally, OGA modulates the -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR), down-regulating GluA2-containing receptors in developing and mature neurons. These findings highlight OGA as a key regulator of excitatory synaptic remodeling and a therapeutic target for synapse-related pathologies such as Alzheimers disease and autism.

neuroscience↗

bioRxiv2025

A Human Angelman Syndrome Class II Pluripotent Stem Cell line with Fluorescent Paternal UBE3A Reporter

IntroductionAngelman Syndrome (AS) is characterized in large part by the loss of functional UBE3A protein in mature neurons. A majority of AS etiologies is linked to deletion of the maternal copy of the UBE3A gene and epigenetic silencing of the paternal copy. A common therapeutic strategy is to unsilence the intact paternal copy thereby restoring UBE3A levels. Identifying novel therapies has been aided by a UBE3A-YFP reporter mouse model. This study presents an analogous fluorescent UBE3A reporter system in human cells. MethodsPreviously derived induced Pluripotent Stem Cells (iPSCs) with a Class II large deletion at the UBE3A locus are used in this study. mGL and eGFP are integrated downstream of the endogenous UBE3A using CRISPR/Cas9. These reporter iPSCs are differentiated into 2D and 3D neural cultures to monitor long-term neuronal maturation. Green fluorescence dynamics are analyzed by immunostaining and flow cytometry. ResultsThe reporter is successfully integrated into the genome and reports paternal UBE3A expression. Fluorescence expression gradually reduces with UBE3A silencing in neurons as they mature. Expression patterns also reflect expected responses to molecules known to reactivate paternal UBE3A. DiscussionThis human-cell-based model can be used to screen novel therapeutic candidates, facilitate tracking of UBE3A expression in time and space, and study human-specific responses. However, its ability to restore UBE3A function cannot be studied using this model. Further research in human cells is needed to engineer systems with functional UBE3A to fully capture the therapeutic capabilities of novel candidates.

cell biology↗