Biology subjects

Lee, S.

Publications and source records attributed to Lee, S..

At least 55 records · Page 3Linked to original sources

bioRxiv2026

Proteomic Profiling Identifies CLDN3 as a Tumor-Selective Therapeutic Target in Small Cell Lung Cancer

Small cell lung cancer (SCLC) remains a highly lethal disease with limited targetable surface antigens beyond delta-like ligand 3 (DLL3). We sought to systematically identify and validate tumor-selective cell-surface targets in relapsed SCLC. To this end, we developed an integrated proteogenomic pipeline combining single-cell RNA sequencing, combinatorial optimization, mass spectrometry-based proteomics, and immunohistochemistry to systematically map the SCLC surfaceome of 49 tumors across 25 patients with relapsed SCLC. Using this approach we found claudin-3 (CLDN3) to be a consistently expressed and tumor-selective antigen, with broader coverage than DLL3, which is the current clinical benchmark. CLDN3 was highly expressed across various treatment states, while maintaining low expression in most nonmalignant tissues. Functional validation using a novel CLDN3-specific monoclonal antibody (ABN501) demonstrated NK cell-mediated cytotoxicity in vitro and tumor regression in vivo, as well as a favorable safety profile. These findings support clinical development of CLDN3-directed therapies and demonstrate the utility of integrated proteogenomic approaches for antigen discovery in solid tumors.

cancer biology↗

bioRxiv2026

Orthogonal CRISPR screens and human liver chimeric mice identify hepatitis B virus host factors

Hepatitis B virus (HBV) chronically infects approximately 250 million people worldwide, and reliable curative therapies are lacking. A broader understanding of viral-host interactions could accelerate efforts to find new host-centric therapeutic targets. However, inefficient cell culture systems and limited replication markers compatible with pooled screening have precluded the widespread use of genetic perturbation screens. Here, we performed the first pooled, genome-wide CRISPR-Cas9 knockout (KO) screen with authentic HBV infection and integrated these results with two orthogonal pooled screens to identify host factors. We selected 72 genes for a multi-step assessment that included arrayed validation assays using both HBV infection and pgRNA transfection. We then independently tested thirteen genes using high-efficiency bulk KO experiments to guide further investigations of both antiviral and proviral factors. In both KO and siRNA-mediated knockdown experiments, depletion of the top antiviral factor, EXOC1, enhanced multiple HBV replication markers, and transcriptomic analysis revealed activation of hypoxia- and HIF-1 gene signatures. Three proviral factors, IRF2, WDR48, and ZCCHC14, were investigated in vivo using a human liver chimeric mouse model, which demonstrated that ZCCHC14 KO greatly reduced HBV replication and spread. Together, these complementary in vitro and in vivo platforms expand the catalog of HBV host factors and provide a scalable framework for host target discovery.

microbiology↗

bioRxiv2026

Programming protein shape as an explicit design layer via CAD blueprint-guided diffusion

In engineering, industrial design principles link geometric form with function. In nature, global protein shape, curvature, and chirality similarly drive critical processes like membrane remodeling and cellular motility. However, explicitly programming these global features remains a challenge for generative AI. Here, we integrate computer-aided design (CAD) blueprints with diffusion models, establishing protein shape as an explicit, programmable layer. We engineered diverse, tunable architectures, including single-chain shapes, scaffolds with unusual twist handedness, and superhelical assemblies, structurally validating 31 of 59 designs via X-ray crystallography or electron microscopy. To demonstrate shape-encoded function, we coupled fluid-flow-optimized helical propellers to an F1-ATPase rotor, yielding a prototype ATP-dependent molecular swimmer. This framework translates industrial design principles to the molecular scale, enabling the programmable engineering of biomimetic architecture with emergent functions. One-sentence summaryIntegrating CAD blueprints with diffusion-based AI translates industrial design principles to the molecular scale, enabling the explicit programming of protein shapes with emergent functions.

bioengineering↗

bioRxiv2026

Rational Redesign of an Fc-Binding Peptide for Multivalent Antibody Assembly

Multivalent antibody assemblies offer opportunities to enhance avidity, organize immune complexes, and modulate higher-order protein interactions, but constructing such architectures from existing immunoglobulin G (IgG) molecules without redesigning the antibody scaffold remains challenging. Here, we report the rational redesign of a Protein A-derived Fc-binding peptide into ADP1, a stable dimeric Fc-binding peptide that directs Fc-mediated antibody assembly. ADP1 was designed from the parent Fc-binding peptide Z34C by preserving the Fc-recognition surface while redesigning the opposite helical surface to promote peptide-peptide association. Biophysical characterization showed that ADP1 retained nanomolar Fc-binding affinity while exhibiting markedly enhanced chemical and proteolytic stability compared with the parent peptide. Structural analyses of ADP1-Fc complexes revealed that ADP1 bridges neighboring Fc regions through a combined ADP1-Fc and ADP1-ADP1 interface, generating spiral higher-order Fc assemblies. This assembly principle was further extended to full-length IgG, where ADP1 promoted higher-order antibody association in a concentration-sensitive manner. In addition, covalent ADP1 functionalization enabled Fc-directed modification of full-length IgG while retaining Fab-mediated antigen recognition, demonstrating the utility of ADP1 as an antibody assembly and functionalization module. Finally, competitive addition of the parent Z34C peptide modulated ADP1-driven antibody assembly, suggesting a potential route for tuning Fc-mediated assembly propagation. Together, this work establishes a redesigned Fc-binding peptide platform for directing multivalent antibody assembly and functionalization without genetic reengineering of the IgG scaffold. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/737354v2_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@57cadeorg.highwire.dtl.DTLVardef@1c7d8e6org.highwire.dtl.DTLVardef@170fb85org.highwire.dtl.DTLVardef@90e6d6_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2026

AI-guided Protein Inhibitor Design for Modulating FAD-dependent Glucose Dehydrogenase Redox Output

Flavin adenine dinucleotide-dependent glucose dehydrogenase (FAD-GDH) is a redox enzyme widely used in glucose monitoring, bioelectronic devices, and enzymatic biofuel cells because of its oxygen-independent catalysis and compatibility with electron-transfer processes. However, protein-based regulators that directly bind GDH and modulate its redox output remain underdeveloped. Here, we present an AI-guided strategy for developing a de novo protein inhibitor targeting FAD-GDH. GDH-targeting candidates generated through structure-based computational design were evaluated by yeast surface display and fluorescence-activated cell sorting, leading to the identification of FAD-GDH inhibitor-1 (FGI-1) as a GDH-targeting inhibitory scaffold. Purified His-MBP-FGI-1 reduced GDH-mediated DCIP reduction, demonstrating attenuation of GDH-derived redox output. Random mutagenesis followed by secondary FACS screening yielded evolved variants with increased GDH-binding signals and enhanced redox-output suppression, showing that the de novo inhibitory scaffold could be functionally tuned through experimental evolution. In addition, an FGI-1-based construct fused to a larger protein module retained GDH-output suppressive activity, and electrode-based measurements showed reduced GDH-derived current output. Because electrode-associated measurements may be influenced by protein-mediated surface shielding and altered electron-transfer accessibility, this decrease was interpreted conservatively as attenuation of GDH-derived electrochemical output rather than direct evidence of active-site inhibition. Together, this work establishes an AI-guided design-validation workflow for developing protein inhibitors that modulate FAD-GDH redox output and provides a foundation for protein-level control of enzyme output in biosensing and bioelectronic applications.

biochemistry↗

bioRxiv2026

Microvascular pathology of proteotoxic endothelial signature characterizes Progressive Supranuclear Palsy

Cerebrovascular pathology is increasingly implicated in neurodegenerative diseases, yet its pathomechanistic contribution remains poorly defined. Building on prior evidence of dysregulated iron and oxygen homeostasis in early-affected brain regions of progressive supranuclear palsy (PSP), we hypothesized that brain microvascular alterations may play an etiological role in select neurodegenerative proteinopathies. First, we conducted a systematic neuropathological evaluation of 178 brains from the University Health Network Neurodegenerative Brain Collection, including Alzheimers disease-related neuropathologic change (ADNC; n=30), Lewy body disease with high or intermediate ADNC (n=38) and low ADNC (n=16), multiple system atrophy (MSA; n=14), PSP (n=39), frontotemporal lobar degeneration with TDP-43 proteinopathy (FTLD-TDP; n=10), and controls (n=31). Arteriolosclerosis, microinfarction, and calcification were assessed in the basal ganglia and frontal cortex. Iron burden was correlated by quantification of Perls staining in MSA and PSP, where vessel pathology was most severe. Single-nucleus RNA-sequencing (snRNA-seq) of frontal cortex tissue from control (n=5) and PSP (n=8) cases with varying arteriolosclerosis severity was performed to characterize the vascular transcriptome, with validation against an independent snRNA-seq evaluation of PSP (n= 11), Picks disease (n=9), AD (n=10), and control (n=10) brains. Histological analysis revealed disease-specific involvement of microvascular pathology in neurodegenerative diseases, identifying PSP to demonstrate most prominent and widespread vessel wall thickening across regions examined. Regression analysis using demographic, APOE and MAPT genetic risk status, and neuropathological features of cases corroborated the distinct association with PSP pathology. Elevated iron load in early affected regions of MSA and PSP brains correlated with greater vessel wall thickening, suggesting a possible pathomechanistic relationship between the two disease physiologies. snRNA-seq analysis of vascular transcriptome identified robust upregulation of heat shock proteins and hypoxia-related genes in PSP endothelial cells and pericytes across both datasets. Importantly, we found the proteotoxic signature to be strongly associated with higher vessel scores in PSP cases, linking microvascular morphology to endothelial dysfunction. Our comprehensive neuropathological evaluation coupled with correlative snRNA-seq analysis establish PSP-specific arteriolar thickening associated with endothelial proteotoxic state as a candidate pathogenic mechanism. The cerebral arteriolar unit represents a compelling therapeutic target for disease modification in PSP.

pathology↗

bioRxiv2026

PanvaR: An R package for fine-mapping and visualizing results from genome-wide association studies

Genome-wide association studies (GWAS) use statistical models to correlate single nucleotide polymorphisms (SNPs) to a phenotype of interest. This scan of the entire genome identifies regions of association with a phenotype, but due to linkage disequilibrium (LD), GWAS on their own cannot identify single genes responsible for phenotypic variation. Rather, fine-mapping of GWAS regions is required, necessitating the use of additional tools and software. With the introduction of more pangenomic resources in a number of crops (Guo et al. 2025; Hufford et al. 2021), the fidelity of these fine-mapping efforts is growing, presenting the opportunity to leverage new information about allelic variation towards gene discovery (Shi et al. 2023; Della Coletta et al. 2021). Panvar is a tool developed to integrate existing software and resources to perform GWAS and fine-mapping in one seamless step. For each identified GWAS peak, panvaR outputs information about LD and SNP effect prediction for each SNP and by layering locations of nearby genes, creates a refined list of possible candidate genes. We have implemented Panvar as an R package, "panvaR", which runs the analysis functions, creates interactive and static visualizations, and outputs results tables. This tool seeks to bridge the gap between GWAS and gene speeding up an important step of quantitative genetic studies.

genomics↗

bioRxiv2026

Developmental stage-dependent decoupling of molecular and phenotypic responses to heat in wheat grain

Heatwaves during flowering and grain development threaten global wheat production, yet the extent to which developmental stage shapes molecular and phenotypic responses remains unclear. Here, we investigated whether heat stress (36/29{degrees}C for 48 h) imposed at closely spaced developmental stages surrounding anthesis generates stage-specific molecular responses that are associated with subsequent effects on grain properties. Heat exposure increased floret abortion most strongly when it was imposed at the trinucleate stage (TN; [~]48%) compared with the binucleate (BN) and early post-anthesis stages. Methylation levels were largely unaffected by prior heat treatments; however, heat induced stage- and locus-specific methylation changes, particularly at BN. Spatial RNA-seq revealed tissue-specific heat responses that were distinctively different in BN and TN. Integrated metabolomic analyses revealed stage-dependent metabolic reprogramming, including shifts towards stress-associated pathways. Despite heat stress at BN generating broader molecular reprogramming in developing grain, exposure at TN produced stronger effects on grain set and composition, revealing a decoupling between the magnitude of responses and phenotypic outcomes. Together, these findings demonstrate that subtle differences in developmental stage influence the complex molecular responses to heat and subsequent grain properties.

plant biology↗

bioRxiv2026

Anticipatory modulation of motor unit discharge rate before rapid isometric elbow flexion force production

Surface electromyography (EMG) studies have demonstrated anticipatory muscle activation prior to predictable voluntary actions. However, the mechanism through which this preparation is expressed, whether through motor-unit recruitment or discharge-rate modulation, remains to be elucidated. We employed surface decomposition EMG to quantify motor-unit behavior prior to self-paced rapid isometric elbow-flexion force pulses. Twelve healthy young men were instructed to generate elbow-flexion pulse force at 30%, 40%, and 50% of the maximal voluntary contraction (MVC) at a self-selected time. Motor-unit activity was decomposed from the biceps brachii and triceps brachii muscles, and the normalized active motor-unit number and mean discharge rate were analyzed prior to pulse onset. In the agonist, the pre-pulse increase mean discharge rate exhibited a higher value than the change in detected motor-unit count, particularly at the 40% and 50% MVC targets. The discharge-rate increase scaled with the target force, with a heightened response observed in high-threshold as compared to low-threshold motor units. Antagonist recordings with sufficient decomposition yield exhibited a similar discharge-rate-dominant pattern; however, these data were available from a smaller sample size. Present findings suggest that discharge-rate modulation is a primary motor-unit-level feature of anticipatory preparation for rapid isometric force production.

neuroscience↗

bioRxiv2026

Inherited proliferation states organize plant transcriptomes

A central question in plant biology is whether natural transcriptome variation primarily reflects environmental adaptation or intrinsic genetic programs. Using transcriptomes from 665 natural Arabidopsis thaliana accessions, we show that inherited population structure, not environmental gradients, is the primary organizer of global transcriptome architecture. Genomic population structure uniquely explains 30.0% of transcriptome-wide variation, while environmental variables account for only 2.5%. This architecture is dominated by a single co-expression program (proliferation Module Eigengene: prolifME), enriched for cell cycle regulation and ribosome biogenesis, regulated by a bipartite transcription factor architecture, and genetically encoded at discrete loci with broad trans-regulatory effects. This proliferation program is associated with reduced plant size, biomass, growth rate and water-use efficiency, and these relationships persist under common-garden conditions. Gene contributions to this proliferation program are non-randomly conserved in rice and maize across 150-200 million years of divergence. These findings identify an intrinsic transcriptional proliferation program as a primary state of plant population transcriptomes, conserved across plant species.

plant biology↗

bioRxiv2026

Low-abundance αSyn-112 promotes αSyn-140 aggregation in vitro and forms immunoreactive deposits in Parkinson's disease brain tissue

The aggregation of -synuclein (Syn) is a molecular hallmark of Parkinsons disease (PD) and other synucleinopathies. Understanding the molecular mechanisms that determine the aggregation of this protein may thus facilitate the development of disease-modifying therapies. While Syn is most commonly expressed as a 140-residue protein (Syn-140), recent evidence suggests an involvement of alternatively spliced Syn isoforms in disease onset and progression. Here, we report and characterise the interaction between Syn-140 and the aggregation-prone Syn-112 variant, one of the most abundant Syn splice isoforms. We found that amounts as low as 1% of Syn-112 accelerate the nucleation and aggregation of Syn-140. To further investigate this phenomenon, we employed MALDI-MS and NMR spectroscopy, confirming that Syn-140 and Syn-112 monomers interact strongly with one another. Furthermore, to assess the association of Syn-112 with disease pathology, we performed immunohistochemical staining combined with confocal microscopy on PD brain samples. Thereby, we found an increase in the number as well as the area of Syn-112 immunoreactive aggregates compared to healthy controls. These results illustrate how low-abundance Syn splice isoforms can modulate the aggregation landscape of Syn-140 and in turn contribute to the molecular heterogeneity of synucleinopathies.

biophysics↗

bioRxiv2026

Targeted depletion of CD38+ autoreactive T cells prevents type 1 diabetes

Type 1 diabetes (T1D) is caused by T cell-mediated autoimmune destruction of insulin-producing islet {beta}-cells. Treatment with T-cell depleting therapies delays the progression of stage 2 and 3 T1D, but these agents exert broad immunosuppressive effects on T cell populations, including T regulatory cells (Tregs), which are key in promoting immune tolerance. We evaluated non-obese diabetic (NOD) mice and recently diagnosed T1D patients and identified CD38 as a marker for pathogenic T cell populations. Using adoptive T-cell transfer in Recombination Activating Gene 1 knockout NOD mice and in a humanized mouse model of autoimmune diabetes, we demonstrated that CD38-expressing autoreactive T cells drive diabetes pathogenesis. Furthermore, we found that selective depletion of CD38+ cells, using an anti-CD38 monoclonal antibody (mAb), prevents insulitis and diabetes onset without depleting CD4+CD25+ Tregs. Administration of anti-CD38 mAb did not adversely affect islet function and may selectively eliminate immunogenic senescent islet {beta}-cells. These results support the strategy of selectively depleting diabetogenic T cells using an anti-CD38 mAb to treat T1D and restore immune tolerance. Therefore, transient depletion of autoreactive T cells using anti-CD38 mAb may provide a novel strategy to prevent or abrogate {beta}-cell autoimmunity in T1D. One sentence summaryPathogenic autoreactive are characterized by CD38 overexpression, and their selective depletion with anti-CD38 monoclonal antibody prevents autoimmune diabetes.

immunology↗

bioRxiv2026

Megalin deficiency perturbs retinal homeostasis and impairs cathepsin D processing and phagosome-lysosome maturation in the retinal pigment epithelium

The multiligand endocytic receptor, megalin (LRP2), is expressed in the retinal pigment epithelium (RPE) and patients lacking the receptor develop high myopia. Despite its established role in retinal development, the contribution of megalin to retinal homeostasis in the normally developed/mature eye remains poorly understood. Here, we investigated megalin function using an inducible knockout mouse (KO) model and human iPSC-derived RPE with megalin knockdown (KD) to distinguish post-developmental homeostatic functions from developmental effects. In vivo, megalin ablation caused progressive retinal degeneration and visual impairment, with morphological abnormalities in the RPE but no changes in myopia-associated ocular phenotypes including axial length and intraocular pressure. Proteomic profiling of megalin-KO RPE revealed reduction of autophagy-related proteins. In line with this, megalin deficiency was associated with accumulation of pro-cathepsin D, and perturbed rhodopsin turnover. This was supported in vitro, where trafficking of photoreceptor outer segment (POS) containing phagosomes to lysosomes was reduced, suggesting disturbed phagosome maturation. Megalin KD did not measurably impair initial uptake of POS discs, but delayed rhodopsin degradation, indicating defective post-ingestion processing. Together, these findings establish megalin as a key regulator of retinal homeostasis in the mature eye by controlling phagosome-lysosome fusion in the RPE and suggest that megalin dysfunction contributes to slowly progressive retinal degeneration. This positions megalin as a potential therapeutic target in lysosomal degenerative diseases in the retina.

Cell Biology↗

bioRxiv2026

A Neuronal Gene Expression Program Underlying Sustained Network Activity

Neurons function within interconnected networks and must continuously adjust their firing properties to maintain stable network activity. Although these adjustments require long-lasting molecular changes, the neuronal gene expression programs that support sustained network activity remain largely unknown. To address this, we developed CalTRAP-seq, a method for profiling gene expression in active neurons via calcium-dependent ribosome tagging. Applying CalTRAP-seq to primary neurons exhibiting synchronous network bursting revealed a gene expression program distinct from stimulus-induced immediate early gene responses, instead enriched for regulators of neuronal excitability. This program was accompanied by widespread alternative splicing of synaptic genes. Notably, neurons participating in network activity exhibited increased nuclear speckle formation, condensates implicated in splicing regulation. Disruption of nuclear speckles impaired synchronous burst dynamics. Together, these findings identify a gene expression program associated with sustained network activity that is complementary to stimulus-responsive gene expression, providing insight into how neurons coordinate gene expression to support stable network function.

neuroscience↗

bioRxiv2026

Individual, behavioural, and environmental determinants of personal light exposure in daily life: A multi-country wearable and experience-sampling study

Light supports circadian regulation and is associated with non-communicable diseases, yet ocular exposure patterns remain poorly understood. We recruited 191 adults across nine sites in seven countries, combining diaries and contextual reports with near-eye (141 participants; 816 participant-days) and complementary chest-level measurements (154 participants; 902 participant-days). In near-eye analyses, only 24.0% of recorded daytime minutes met the recommendation of at least 250 lx melanopic equivalent daylight illuminance; 63.3% of pre-sleep and 87.7% of bedside sleep-environment minutes met their respective limits. Variation among people and days within sites exceeded that among sites (ratio 1.99; 95%-CI, 1.29-4.77). In hourly models, light source and setting contributed the largest shares of fitted variation after time of day. Site-average outdoor exposure was 9.35 times that while awake at home (95%-CI, 6.95-12.59; 714.2 versus 76.4 lx). This baseline identifies a daytime exposure gap and supports testing interventions in everyday settings to improve light exposure and health.

physiology↗

bioRxiv2026

Kinetics of Hypoglycemia in Diabetes Patients Informs Development of New Modes of Glucagon Therapy

Insulin therapy revolutionized the care of patients with diabetes starting [~]100 years ago, yet insulin-induced hypoglycemia remains a serious life-threatening complication of insulin therapy. Glucagon is a highly effective treatment; however current dosage forms remain under-utilized due to poor patient compliance. The development of improved and situation-specific glucagon therapies remains challenging due to the poor drug stability and incomplete knowledge of the kinetics of different hypoglycemic events. Thus, we analyzed continuous glucose monitor (CGM) data from 1135 patients with type 1 diabetes (T1D) representing 246.18 patient years. We show that a surprisingly large proportion of hypoglycemic episodes (20-30%) are follow-on events resulting from under-treatment of prior events, and that the average duration of independent hypoglycemic events can last up to 79 to 108 minutes. We further show that the kinetics of hypoglycemic onset and persistence varies significantly by patient history, severity, time of occurrence. Guided by these findings, we recognize the opportunity to develop high-density, readily-soluble, and thermostable (ReST) solid glucagon formulations, and painless application-specific microneedle-patches that are in line with the timing needs of T1D patients who are awake and asleep. Thus, we demonstrate (1) on-demand patches for rapid prevention or treatment of mild hypoglycemia during the day, and (2) enzyme-driven hypoglycemia-responsive patches supporting autonomous glucagon release during the night. We show excellent in vitro glucagon stability, loading, and release kinetics of both systems and demonstrate their ability to treat hypoglycemia in diabetic animals. The engineering of these delivery systems demonstrates the potential of human CGM data and solid glucagon formulations to enable new modes of glucagon therapy, thereby expanding the clinical role of glucagon beyond the emergency setting.

bioengineering↗

bioRxiv2026

Beta-Adrenergic Stimulation and MYH7 G256E Mutant Gene Dosage Drive Hypertrophic Cardiomyopathy Phenotype Penetrance

AimsHypertrophic cardiomyopathy (HCM) is the most prevalent genetic heart disorder, characterized by significant phenotypic variability even among individuals with identical MYH7 mutations. This study aims to elucidate factors contributing to this variability and identify drivers of phenotype penetrance. We compared the baseline phenotypes of a highly penetrant MYH7 H251N mutation and the variably penetrant MYH7 G256E mutation and investigated the impact of adding beta-adrenergic stimulation and homozygosity on disease phenotype penetrance using cardiomyocytes from an isogenic line of human induced pluripotent stem cells (hiPSC-CMs). Methods and ResultsIsogenic hiPSCs with MYH7 H251N and MYH7 G256E mutations were generated using CRISPR/Cas9 technology and differentiated into cardiomyocytes (CMs). Single-cell RNA sequencing (scRNAseq) and functional analysis of contractile function revealed consistent HCM phenotype presentation in H251N CMs, whereas G256E CMs exhibited a subtle and more variable phenotype. Beta-adrenergic stimulation induced a distinct metabolic stress response in G256E CMs, characterized by impaired mitochondrial ATP upregulation. Increasing mutant gene dosage from hetero- to homozygosity led to consistent increase in hypertrophic and structural gene expression changes in G256E CMs at RNA and protein levels. These changes were distinct from the changes observed with stress response. Importantly, homozygous G256E CMs exhibited a hypercontractile functional and disorganized structural phenotype. Across multiple experimental conditions, we identified consistent increase in cardiomyocyte specific transcriptomic markers such as NPPB, APOE, PDLIM3 and ANKRD1. ConclusionsOur study highlights the use of a variably penetrant MYH7 mutation to investigate factors that influence HCM phenotype penetrance. Specifically, we found that mutant gene dosage and beta-adrenergic stimulation induce distinct HCM disease phenotypes, providing novel insights into mechanisms that may contribute to variable disease expression in HCM. Translational PerspectiveHCM is characterized by significant phenotypic variability, complicating both diagnosis and clinical management. This study explores the factors driving HCM phenotype penetrance using isogenic hiPSC-CMs with MYH7 mutations. We demonstrate that beta-adrenergic stimulation and increased mutant gene dosage significantly impact HCM disease penetrance. Beta-adrenergic stimulation triggers metabolic stress responses, while increased gene dosage leads to a hypercontractile and structurally disorganized phenotype. These findings provide insight into how specific modifiers can shape disease-associated phenotypes in HCM model systems. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/729411v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@16a72d7org.highwire.dtl.DTLVardef@1999ca6org.highwire.dtl.DTLVardef@1fb8752org.highwire.dtl.DTLVardef@7148b2_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2026

TDKC (Target Distilled K-mer Classifier): Ultrafast and Memory-Efficient Sequence Classification for Target Pathogen Diagnostics

Metagenomic sequencing can identify pathogens from clinical samples without prior knowledge of the causative agent. Yet, as sequencing workflows scale to process thousands of multiplexed samples simultaneously, classifying these samples against massive reference databases creates a significant computational bottleneck. Furthermore, large-scale applications such as screening public sequence repositories remain computationally challenging. Existing metagenomic classifiers are designed for full-taxon classification, where the goal is to identify all organisms in a sample. However, many diagnostic applications focus on detecting a specific set of clinically relevant pathogens. This constraint can be exploited to significantly lower computational costs. Here we present TDKC (Target Distilled K-mer Classifier), a method for targeted metagenomic classification. TDKC constructs a compact index by distilling target-specific k-mers from a full-taxon reference database. When classifying clinical samples, TDKC uses 16.9-33.6x less memory and is 5.1-34.7x faster than per-read full-taxon and targeted classifiers (Kraken2, Centrifuger, CLARK), while maintaining high sensitivity and low false positive rates. Against the sketch-based profiler Sylph, TDKC remains 3.8x faster and uses 8.7x less memory. TDKC also supports per-k-mer accession tracking across over 3 million source accessions for downstream subtype analysis, and domain-level detection of bacteria, archaea, and viruses. By reducing the index to only the pathogens of interest, TDKC makes targeted pathogen detection feasible at scale.

bioinformatics↗