Biology subjects

Nam, J.

Publications and source records attributed to Nam, J..

16 recordsLinked to original sources

bioRxiv2026

Decoding and Targeting Coordinated CDKN1A and CDKN2A Senescence Programs in ECM-Dominant Cardiovascular Pathologies

Cellular senescence is a hallmark of aging and an emerging therapeutic target; however, its role as a context-specific driver of disease remains incompletely defined, and senolytic therapies have shown inconsistent clinical benefit. Here, we identify extracellular matrix (ECM)-dominant pathologies as a major class of senescence-driven disease, characterized by inflammation, matrix degeneration, and progressive tissue dysfunction. Using integrated single-cell transcriptomics, spatial profiling, and multiplex imaging across human specimens and murine models, we demonstrate that senescent fibroblasts, rather than canonical myofibroblasts, constitute the principal disease-driving cell state in myxomatous mitral valve disease (MMVD) and related conditions. These cells exhibit coordinated CDKN1A inflammatory and CDKN2A ECM-remodeling programs that form a feed-forward circuit linking immune activation to matrix disorganization and functional decline. Senescence extends beyond fibroblasts to endothelial and immune compartments, establishing a multicellular senescent milieu that reinforces intercellular crosstalk and disease progression. Senolytic treatment (dasatinib plus quercetin or fisetin) restores ECM architecture and improves cardiac function, outperforming pathway-specific anti-inflammatory and antifibrotic approaches. Cross-disease analyses further reveal conservation of this coordinated CDKN1A/CDKN2A senescence programs across multiple ECM-dominant cardiovascular diseases, including aortic aneurysm and calcific valve disease. Notably, in vivo single-cell transcriptomic profiling following multiple senolytic treatments provides whole-transcriptome resolution of context-dependent cellular responses. Collectively, these findings establish context-specific senescence as a central organizing mechanism in ECM-dominant diseases and support a shift from generalized anti-aging strategies toward precision senolytic prevention or therapy. Given that valvular and aortic diseases affect millions and increase markedly with age to a prevalence comparable to major cancers, these results indicate a potential solution to a substantial and underrecognized clinical burden.

Cell Biology↗

bioRxiv2026

Multi-omic profiling of human and mouse dorsal root ganglia enables targeted gene delivery to nociceptors

Chronic pain conditions are often driven by hyperexcitability of nociceptors, the peripheral sensory neurons that detect noxious stimuli. Recent single-cell transcriptomic studies have begun to clarify the molecular identity of distinct peripheral sensory neuron subtypes, but tools that restrict transgene expression to nociceptors while sparing other dorsal root ganglion (DRG) subtypes remain limited. Here, we combined single-nucleus multi-omic profiling of human and mouse DRG with in vivo AAV enhancer screening to identify cis-regulatory elements that drive biased AAV expression in mouse DRG nociceptors and human iPSC-derived nociceptors. We then validated that an enhancer AAV designed to express Kir2.1 preferentially in nociceptors reduces DRG neuronal excitability. Leveraging these multi-omic datasets, we trained a sequence-based model to decode the cis-regulatory logic of nociceptors, enabling both the prioritization of native candidate elements and the design of synthetic enhancers with a range of nociceptor targeting properties. Together, these cross-species multi-omic resources define conserved DRG regulatory programs and provide a viral toolkit for pain research with potential translational applications for patients with refractory pain.

neuroscience↗

bioRxiv2026

Engineering an in vitro model of demyelinated spinal cord tissue

Demyelinating diseases are a group of complex neurodegenerative disorders characterized by damage to the myelin, the protective sheath that insulates and supports efficient nerve signal conduction. Such a loss of myelin causes the formation of lesions not only in the brain but also often in the spinal cord (SC). Despite the high prevalence of SC lesions among patients, existing models mostly focus on those in the brain, inadequately capture the unique anatomical and physiological features of SC pathology. In this study, we developed a robust, reproducible in vitro model of SC demyelination by combining microwell technology and piezoelectric scaffolds to engineer human neural stem cell (hNSC)-derived nerve tissues featuring aligned, myelinated, extended axons up to 2000 {micro}m in length. We utilized distinct chemical treatments to induce demyelination with or without axonal degeneration: a cuprizone cocktail, a copper chelator combined with inflammatory cytokines, and lysophosphatidylcholine (LPC). Electrophysiological assessments validated the physiological relevance of our model, demonstrating impaired signal transmission and neural connectivity akin to in vivo demyelination pathology. Our versatile platform thus provides a valuable tool for elucidating SC demyelination pathophysiology and exploring potential therapeutic interventions.

bioengineering↗

bioRxiv2025

Non-invasive peripheral delivery of CDNF fragment protects neurons in models of Parkinsons and ALS

Non-invasive delivery of brain therapeutics is a key challenge for treating neurodegenerative diseases. Here, we discovered a novel carboxy (C)-terminal fragment of cerebral dopamine neurotrophic factor (C-CDNF) that protects dopamine (DA) and motoneurons (MNs) in rodent models of Parkinsons disease (PD) and amyotrophic lateral sclerosis (ALS). C-CDNF retains the same structure as CDNF and similarly to CDNF regulates cell stress pathways but unorthodoxly enters cultured neurons and passes through the blood-brain barrier. In vivo, intracranially or peripherally delivered C-CDNF improves motor deficits, protects DA neurons, and restores motor behavior in a rat model of PD. Subcutaneous C-CDNF also protects MNs and reduces microglial activation in an ALS model. Based on our findings, beginning C-CDNF treatment soon after diagnosis is anticipated to delay progression of PD and ALS, thereby improving treatment outcome. Thus, systemic delivery of C-CDNF should simplify the administration of protein-based therapeutics to patients while reducing treatment risk and financial burden for patients and families.

neuroscience↗

bioRxiv2024

Impact of a transient neonatal visual deprivation on the development of the ventral occipito-temporal cortex in humans

How does experience shape the development of visual brain regions? We demonstrate that a transient period of visual deprivation early in life in humans leads to permanent alteration in the function of the early visual cortex (EVC), while leaving the categorical coding of downstream ventral occipito-temporal cortex (VOTC) mostly unaffected. We used fMRI to characterize the brain response to five categories (faces, bodies, objects, buildings and words) in a rare group of cataract-reversal individuals who experienced a transient period of blindness early in life, and in matched participants with typical visual development. We show that the encoding of low-level visual properties is impaired in EVC in cataract-reversal participants, while the categorical response in the VOTC is preserved. When altering the visual properties of our stimuli to mimic in controls the deficit of EVC response of the cataract, we observe a cascading alteration of the categorical coding from EVC to VOTC that is not observed in the cataract-reversal group. Our results show that a typical visual categorical organization develops in VOTC despite a period of visual deprivation early in life and in the presence of impaired processing in EVC. A deep neural network exposed to a visual diet that emulates the experience of our participants reproduced our brain observations within an artificial system. These findings emphasize how specific visual regions are differently affected by a period of visual deprivation early in life, with information encoded in EVC being permanently affected, while the categorical coding in VOTC shows resilience to deprivation.

neuroscience↗

bioRxiv2024

Integration of Imaging-based and Sequencing-based Spatial Omics Mapping on the Same Tissue Section via DBiTplus

Spatially mapping the transcriptome and proteome in the same tissue section can significantly advance our understanding of heterogeneous cellular processes and connect cell type to function. Here, we present Deterministic Barcoding in Tissue sequencing plus (DBiTplus), an integrative multi-modality spatial omics approach that combines sequencing-based spatial transcriptomics and image-based spatial protein profiling on the same tissue section to enable both single-cell resolution cell typing and genome-scale interrogation of biological pathways. DBiTplus begins with in situ reverse transcription for cDNA synthesis, microfluidic delivery of DNA oligos for spatial barcoding, retrieval of barcoded cDNA using RNaseH, an enzyme that selectively degrades RNA in an RNA-DNA hybrid, preserving the intact tissue section for high-plex protein imaging with CODEX. We developed computational pipelines to register data from two distinct modalities. Performing both DBiT-seq and CODEX on the same tissue slide enables accurate cell typing in each spatial transcriptome spot and subsequently image-guided decomposition to generate single-cell resolved spatial transcriptome atlases. DBiTplus was applied to mouse embryos with limited protein markers but still demonstrated excellent integration for single-cell transcriptome decomposition, to normal human lymph nodes with high-plex protein profiling to yield a single-cell spatial transcriptome map, and to human lymphoma FFPE tissue to explore the mechanisms of lymphomagenesis and progression. DBiTplusCODEX is a unified workflow including integrative experimental procedure and computational innovation for spatially resolved single-cell atlasing and exploration of biological pathways cell-by-cell at genome-scale.

systems biology↗

bioRxiv2024

All-Optical Multimodal Mapping of Single Cell Type-Specific Metabolic Activities via REDCAT

Metabolism underlies cell growth, survival, and function, yet its activities vary widely across cell types and tissue environments. Spatially resolving these processes in situ at single-cell resolution is essential to advance our understanding of cellular function and tissue physiology in health and disease. However, existing approaches are limited by either destructive workflows, insufficient spatial resolution and biochemical specificity, or lack of direct linkage to cell identity. Here, we present Raman Enhanced Delineation of Cell Atlases in Tissues (REDCAT), a multimodal all-optical platform that integrates stimulated Raman scattering, autofluorescence redox imaging, second harmonic generation, and high-plex immunofluorescence to co-map metabolic activities and cell types within the same tissue section. REDCAT achieves subcellular resolution profiling of protein, lipid, redox, and nuclear acid metabolism, together with extracellular matrix composition, in both FFPE and fresh-frozen human tissues. Applied to normal lymph nodes, REDCAT delineated distinct redox and lipid remodeling programs across germinal center B-cell zones and immune subsets, highlighting cell-type-specific metabolic specialization. In lymphoma, it revealed profound metabolic reprogramming, including extensive lipid accumulation, nuclear metabolic heterogeneity, and a transitional metabolic state associated with transformation from chronic lymphocytic leukemia to diffuse large B-cell lymphoma, thereby illuminating tumor evolution in situ. In human liver, REDCAT resolved cell-type-specific lipid droplet diversity and zonation-dependent nuclear metabolic gradients, uncovering new principles of spatial metabolic organization. By directly linking cell identity with spatial metabolic states at single-cell or subcellular resolution, REDCAT establishes a broadly applicable framework for studying immune function, tumor progression, and tissue physiology, and offers a new path to deciphering the metabolic basis of health and disease.

cell biology↗

bioRxiv2024

Accurate antibody loop structure prediction enables zero-shot design of target-specific antibodies

Protein loops, characterized by their versatile structures with varying sizes and shapes, can recognize a wide range of targets with high specificity and affinity. The variable loops of the antibody complementarity-determining region (CDR) are particularly crucial for immune responses and therapeutic applications due to their effective target recognition capabilities. Accurate structure prediction of these antibody loops is essential for the efficient in silico design of target-binding antibodies for therapeutic or industrial use. However, predicting antibody loop structures is challenging due to the lack of evolutionary information from related proteins. Thus, a successful ab initio structure prediction method, which operates without structural templates or related sequences, is crucial for the effective design of antibody loop-mediated interactions. This study demonstrates that highly accurate antibody loop structure prediction enables the effective zero-shot design of target-binding antibody loops. The performance of loop design has been shown to depend on the accuracy of ab initio loop structure prediction, as tested with two versions of our design model. The high affinity, diversity, novelty, and specificity of the antibody loops designed with these new methods were validated experimentally on four target proteins.

bioengineering↗

bioRxiv2023

Small molecule activation of m6A mRNA methylation as a novel approach for neuroprotection

N6-Methyladenosine (m6A) is the most common mRNA base modification in eukaryotes. Methylation of adenosine residues to m6A contributes to the regulation of splicing, transport, stability, and translation of mRNA and two main classes of enzymes regulate it. The formation of m6A is catalysed by a methyltransferase complex containing methyltransferase-like 3 (METTL3), METTL14, and Wilms tumour 1-associated protein (WTAP) as well as monomeric METTL16. Demethylation of m6A is catalysed by the fat mass and obesity-associated protein FTO and the RNA demethylase AlkB homolog 5 (ALKBH5). The m6A mRNA methylation dysregulation occurs in the nervous system and in Parkinsons disease (PD), but it remains poorly studied. Moreover, the role of m6A mRNA methylation in neuronal survival, neuroprotection, and neuroregeneration is unclear. We have earlier used high-throughput virtual screening of large compound libraries and identified four unique small-molecule ligands that activate m6A mRNA methylation by binding to the METTL3/14/WTAP complex and enhancing the binding of the methylation substrate SAM to nanomolar concentrations. Following this, we now discovered that two methyltransferase activators at 10 nM concentrations supported the survival and protected dopamine (DA) neurons in culture in growth factor deprivation and 6-hydroxydopamine (6-OHDA) neurotoxin models. In contrast, METTL3/14 inhibitor STM2457 triggered death of DA neurons. For clinical translation we also tested the most efficient compound C4 on induced pluripotent stem cell-derived human DA neurons and in animal model of Parkinsons disease (PD). C4 compound protected human DA neurons from 6-OHDA-induced cell death and increased neurite outgrowth and the number of processes demonstrating that it has both neuroprotective and neurorestorative properties. METTL3/14 activator C4 improved motor behaviour and protected DA neurons and their fibres faster and much more efficiently than GDNF in the rat 6-OHDA model of PD. These are the first specific activators of METTL3/14/WTAP and first demonstration that m6A regulators can protect and regenerate neurons. These data demonstrate that m6A mRNA methylation is a novel pathway regulating neuronal survival and regeneration.

neuroscience↗

bioRxiv2023

Enhanced Peripheral Nerve Regeneration by Mechano-electrical Stimulation

To address limitations in current approaches for treating large peripheral nerve defects, this study evaluated the efficacy of functional material-mediated physical stimuli on peripheral nerve regeneration. Electrospun piezoelectric poly(vinylidene fluoride-trifluoroethylene) nanofibers were utilized to deliver mechanical actuation-activated electrical stimulation to nerve cells/tissues in a non-invasive manner. Using morphologically and piezoelectrically optimized nanofibers for neurite extension and Schwann cell maturation based on in vitro experiments, piezoelectric nerve conduits were implanted in a rat sciatic nerve transection model to bridge a critical-sized sciatic nerve defect (15 mm). A therapeutic shockwave system was utilized to activate the piezoelectric effect of the implanted nerve conduit on demand. The piezoelectric nerve conduit-mediated mechano-electrical stimulation induced enhanced peripheral nerve regeneration, resulting in full axon reconnection with myelin regeneration from the proximal to the distal ends over the critical-sized nerve gap. Furthermore, superior functional recovery was observed by walking track analysis and polarization-sensitive optical coherence tomography, demonstrating the excellent efficacy of the mechano-electrical stimulation strategy for treating peripheral nerve injuries.

bioengineering↗

bioRxiv2023

Synthetic lipid rafts formed by cholesterol nano-patch induce T cell activation

Lipid rafts cluster at the immunological synapse during activation and serve as signaling hubs in T cells. While cholesterol plays a crucial role in lipid raft formation, the impact of their spatial configuration remains less understood. Here, we introduce programmable DNA origami, cholesterol nano-patch, for nanoscale spatial control of cholesterols on live T cell membranes, enabling us to elucidate their roles in lipid raft formation, receptor activation, and intracellular signaling. We demonstrate that CNPs with high-density cholesterol arrangements efficiently bind to the T cell plasma membrane and form a large and polarized coalescence, leading to the stabilization of ordered lipid membrane domains by increasing the local concentration of cholesterols. These synthetic lipid rafts colocalize with flotillin-1, a raft-associated protein, and promote the membrane reorganization and physical segregation of key signaling molecules, such as T cell receptors, Lck and LAT kinases, and CD45. Consequently, this leads to early T cell activation in the absence of antigenic stimulation. Our DNA origami approach demonstrates the direct role of cholesterol in lipid raft formation, leading to membrane phase separation and protein reorganization, which are sufficient to induce early T cell activation.

bioengineering↗

bioRxiv2022

Actin cytoskeleton signaling via MRTF/SRF entrains circadian clock

The circadian clock is entrained to daily environmental cues. Integrin-linked intracellular signaling via actin cytoskeleton dynamics transduces extracellular matrix interactions to Myocardin-related Transcription Factor (MRTF)/Serum Response Factor (SRF)-mediated transcription. Actin cytoskeleton organization in liver displays diurnal oscillations and SRF-MRTF activity exert transcriptional control to entrain clock. By interrogating disparate upstream events involved in actin cytoskeleton-MRTF-A/SRF signaling cascade, here we show that this signaling cascade transduce cellular niche cues to modulate circadian clock function. Pharmacological inhibitions of MRTF-A/SRF, by disrupting actin polymerization or blocking ROCK kinase, induced period lengthening with augmented clock amplitude, and genetic loss-of-functions of Srf or Mrtf-a mimic that of actin-depolymerizing agents. In contrast, actin-polymerization induced by Jasplakinolide shortened period with attenuated amplitude. In addition, interfering with cell-matrix interaction through blockade of integrin, inhibition of focal adhesion kinase or attenuating matrix rigidity led to reduced period length while enhancing amplitude. Mechanistically, we identify that core clock repressors, Per2, Nr1d1, and Nfil3, are direct transcriptional targets of MRTF-A/SRF in mediating actin dynamic-induced clock response. Collectively, our findings defined an integrin-actin cytoskeleton-MRTF/SRF pathway in linking clock entrainment with extracellular microenvironment that may facilitate cellular adaptation to its physical niche. Summary statementOur study revealed the role of actin cytoskeleton-MRTF/SRF signaling in entraining circadian clock to its extracellular physical niche environment.

cell biology↗

bioRxiv2021

Integrin signaling via actin cytoskeleton activates MRTF/SRF to entrain circadian clock

The circadian clock is entrained to daily environmental cues. Integrin-linked intracellular signaling via actin cytoskeleton dynamics transduces cellular niche signals to induce Myocardin-related Transcription Factor (MRTF)/Serum Response Factor (SRF)-mediated transcription. So far, how the integrin-associated signaling cascade may transmit cellular physical cues to entrain circadian clock remains to be defined. Using combined pharmacological and genetic approaches, here we show that the transcription factors mediating integrin to actin cytoskeleton signaling, MRTF-A and SRF, exert direct transcriptional control of core clock components, and that this signaling cascade modulates key properties of clock circadian activity. Pharmacological inhibition of MRTF/SRF activity by disrupting actin polymerization significantly augmented clock amplitude with period shortening, whereas an actin polymerizing compound attenuated oscillation amplitude with period lengthening. Genetic loss-of-function of Srf or Mrtf mimics that of actin-depolymerizing agents, validating the role of actin dynamics in driving clock function. Furthermore, integrin-mediated focal adhesion with extracellular matrix and its downstream signaling modulates the circadian clock, as blockade of integrin, focal adhesion kinase or Rho-associated kinase (ROCK) increased clock amplitude and shortened period length. Mechanistically, we identify specific core clock transcription regulators, Per1, Per2 and Nr1d1, as direct target genes of MRTF-A/SRF. Collectively, our findings uncovered an integrin-actin cytoskeleton-MRTF/SRF signaling cascade in linking clock entrainment to its extracellular microenvironment, which may mediate cellular adaptation to its physical niche. Author SummaryThe circadian clock anticipates and adapts to environmental changes. Interestingly, serum, as a universal clock synchronizing signal, drives intracellular actin cytoskeleton reorganization through modulation of MRTF/SRF activity. However, mechanisms that may transduce extracellular niche signals to circadian clock remains to be defined. We hypothesize that integrin-mediated intracellular signaling to actin cytoskeleton links extracellular microenvironment with MRTF/SRF transcriptional regulation to control clock function. Using small molecules and genetic approaches targeting distinct steps of integrin-actin cytoskeleton-MRTF/SRF signaling cascade, we uncover the effects of this pathway in controlling circadian clock oscillation. We also identify specific core clock regulators as direct gene target genes of MRTF and SRF-mediated transcriptional control. Our study revealed how integrin-mediated cellular interaction with its physical environment influences its intrinsic clock properties through signaling transduction via actin cytoskeleton remodeling, and that this mechanism may facilitate circadian clock adaptation to cellular physical niche.

cell biology↗

bioRxiv2020

MANF regulates unfolded protein response and neuronal survival through its ER-located receptor IRE1α

Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an endoplasmic reticulum (ER)-located protein with cytoprotective effects in numerous cell types in vitro and in models of neurodegeneration and diabetes in vivo. So far, the exact mode of its action has remained elusive and plasma membrane or ER-located receptors of MANF have not been identified. We have found that MANF can directly interact with transmembrane unfolded protein response (UPR) receptor IRE1 and compete with the major ER chaperone BiP (GRP78) for the interaction with IRE1. With lower affinities MANF can also interact with other UPR receptors, PERK and ATF6. Using molecular modeling and mutagenesis analysis, we have identified the exact structural MANF regions involved in its binding to the luminal domain of IRE1. MANF attenuates UPR signaling by decreasing IRE1 oligomerization and IRE1 phosphorylation. MANF mutant deficient in IRE1 binding cannot regulate IRE1 oligomerization and fails to protect neurons from ER stress induced death. Importantly, we found that MANF-IRE1 interaction is also crucial for the survival promoting action of MANF for dopamine neurons in an animal model of Parkinsons disease. Our data reveal a novel mechanism of IRE1 regulation during ER stress and demonstrate the intracellular mode of action of MANF as a modulator of UPR and neuronal cell survival through the direct interaction with IRE1 and regulation of its activity. Furthermore, our data explain why MANF in contrast to other growth factors has no effects on naive cells and rescues only ER stressed or injured cells.

cell biology↗

bioRxiv2020

CDNF rescues motor neurons in three animal models of ALS by targeting ER stress

The role of chronic endoplasmic reticulum (ER) stress in the pathophysiology of Amyotrophic lateral sclerosis (ALS), as well as a potential drug target, has received increasing attention. Here, we investigated the mode of action and therapeutic effect of the ER resident protein cerebral dopamine neurotrophic factor (CDNF) in preclinical models of ALS harboring different genetic mutations. We identify that intracerebroventricular (i.c.v.) administration of CDNF significantly halts the progression of the disease and improves motor behavior in TDP43-M337V and SOD1-G93A rodent models of ALS. CDNF rescues motor neurons (MNs) in vitro and in vivo from ER stress associated cell death and its beneficial effect is independent of genetic disease etiology. Notably, CDNF regulates the unfolded protein response (UPR) initiated by transducers IRE1, PERK, and ATF6, thereby enhancing MN survival. Thus, CDNF holds great promise for the design of new rational treatments for ALS.

neuroscience↗

bioRxiv2018

Unbiased genome-scale identification of cis-regulatory modules in the human genome by GRAMc

Although significant advances have been made toward functionally identifying human regulatory elements, existing genome-scale reporter methods preferentially detect either enhancers or promoters. Here we develop GRAMc, a highly reproducible unbiased Genome-scale Reporter Assay Method for cis-regulatory modules (CRMs). GRAMc combines the versatility of traditional reporter constructs and the scalability of DNA barcode reporters, and unites the complementary advantages of several currently available high-throughput reporter assays. We demonstrate that GRAMc can reliably measure cis-regulatory activity of nearly 90% of the human genome in 200 million HepG2 cells with randomly fragmented ~800bp inserts. By using the GRAMc-identified CRMs, we show that CRMs identified in one cell type are useful for predicting gene regulatory programs not only within that cell type but also between cell types or conditions separated in time and space. In addition, the GRAMc-identified CRMs support the hypothesis that SINE/Alu elements are rich sources of regulatory evolution. Finally, the observation that the majority of experimentally identified regulatory elements do not overlap with computationally predicted elements underscores the necessity of an efficient and unbiased genome-scale reporter assay.

genomics↗