Biology subjects

King, A.

Publications and source records attributed to King, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Staphylococcus aureus GpsB, Stk, and VraSR form a cell wall surveillance network to maintain cell envelope integrity and mediate antibiotic resistance

Antibiotic resistance is an alarming threat to public health worldwide. Specifically, drug-resistant Staphylococcus aureus remains a major contributor to global mortality. Therefore, in-depth understanding of antibiotic resistance mechanisms is urgently needed to inform alternative therapeutic strategies. We previously showed that GpsB is a critical regulator of cell wall synthesis in S. aureus. GpsB executes its regulatory role via protein-protein interactions. Yet, how GpsB function is modulated in this organism has remained unclear. Intriguingly, a prior proteomics report indicated that GpsB is phosphorylated at six different residues by the sole serine/threonine kinase Stk. Hence, we explored the physiological significance of GpsB phosphorylation. We identify three phosphosites that are each independently sufficient to drive GpsB phosphoregulation, which likely alters its partner preference. We find that cells lacking gpsB exhibit heightened fosfomycin sensitivity, and that this is exacerbated by stk deletion. Our findings reveal that GpsB collaborates with Stk and the VraSR cell wall stress response system to coordinate the demand-driven availability of cell wall precursors. Collectively, this report establishes GpsB as a pivotal peptidoglycan supply chain sentinel that together with Stk and VraSR forms an integrated cell wall stress monitoring network that maintains cell envelope homeostasis and aids in antibiotic resistance.

microbiology↗

bioRxiv2026

Targeted single-nucleus sequencing of 39,800 neurons reveals extensive low-frequency somatic variants

Recent evidence implicates somatic mutations in disease-associated genes across multiple neurodegenerative disorders, including the identification of two somatic TARDBP variants as a putative cause of frontotemporal lobar degeneration with TDP-43 pathology type C (FTLD-TDP type C). However, these findings remain anecdotal. We performed single-nucleus amplicon sequencing of 39,800 neurons from the superior temporal gyrus of sporadic FTLD-TDP type C patients (34,738 neurons, 52 individuals) and non-demented controls (5,062 neurons, 26 individuals) using the Mission Bio Tapestri platform and investigated somatic variants in TARDBP and other FTLD-TDP-associated genes in a case-control setting. We uncovered an extensive landscape of ultra-low-frequency (<1%) somatic mutations across all analysed genes, including TARDBP. Rare somatic occurrences of known ALS/FTD-associated germline TARDBP variants, including p.N267S and p.M337V, were identified exclusively in patients. Among all targeted genes, TARDBP had the highest proportion of neurons harbouring a somatic variant, and this mutational burden was lower in individuals who died at an older age. Strikingly, the C-terminal domain of TDP-43, where most ALS/FTD germline pathogenic variants reside, exhibited a lower neuronal mutational burden than its other domains. This suggests that neurons harbouring damaging mutations may undergoprogressive loss prior to autopsy, leaving only rare survivors at frequencies too low to detect by bulk or small-scale single-cell approaches. Together, our findings provide an unprecedented resolution into the somatic mutational landscape of post-mitotic neurons in FTLD-TDP type C and reveal a gene-specific layer of neuronal mosaicism that current single-cell WGS studies fail to detect.

genetics↗

bioRxiv2026

Magnesium induces iron starvation and metabolic rewiring to support the viability of cell envelope mutants and antibiotic-stressed cells

Magnesium supplementation permits deletion of otherwise essential genes involved in cell envelope biogenesis in the Gram-positive model bacterium Bacillus subtilis. Yet, the specific underlying mechanism has remained elusive. To address this key knowledge gap, we made use of a mutant lacking ezrA and gpsB. Deletion of both of these genes involved in cell wall synthesis leads to severe growth inhibition which is ameliorated by magnesium addition. Our results indicate that, in the absence of magnesium, this mutant contains elevated levels of labile iron, is impaired in activating the oxidative stress response, and displays extreme sensitivity to iron and manganese intoxication. Intriguingly, we find that an ezrA single deletion, but not gpsB, exhibits heightened susceptibility to excess iron and manganese. This observation allowed us to investigate the source of toxicity and how EzrA may support metal homeostasis. Our data suggests that the major contributor of ROS is the electron transport system involved in cellular respiration. Both genetic and chemical means to reprogram the cells in favor of fermentation alleviate the metal toxicity in cells lacking ezrA. Collectively, our data shows that magnesium limits iron availability and redirects metabolism towards pathways that are preferred during iron scarcity. Consequently, these mechanisms result in reduced ROS production and oxidative stress mitigation. This explains why magnesium supplementation may render essential genes dispensable. In support of this model, we find that addition of magnesium helps cells to circumvent lysis typically caused by the treatment of an antibiotic that disrupts cell wall synthesis. Taken together, our results suggest that unmitigated oxidative stress fueled by labile iron is likely responsible for the detrimental effects of specific gene disruptions and certain antibiotic treatments. By reducing the pool of free iron and reprogramming cellular metabolism, magnesium mitigates oxidative damage and protects cells from ROS-mediated death.

microbiology↗

bioRxiv2026

Practice-dependent refinement of motor execution is retained and broadly transferable but constrained by movement direction

Practice enhances motor acuity, enabling movement execution with greater speed and accuracy. However, the learning principles underlying improvements in speed, accuracy, and efficiency remain less understood than those supporting motor skill acquisition and adaptation. Here, we examined motor execution in a skill-based practice task to characterize learning, retention, and generalization of motor acuity. Using a gamified two-dimensional racing task, right-handed participants controlled a stylus-driven car along a curved track as quickly and accurately as possible. Across two studies (N = 83 total, 54 females), participants completed 300 training laps on Session 1 and returned for Session 2 to assess retention and generalization to novel track configurations: one with altered spatial configuration (rotated track) and one requiring movement in the opposite direction of training (reverse track). Movement speed improved rapidly and showed robust, though incomplete, retention across sessions. Speed improvements generalized substantially to both novel tracks. Accuracy was high at training onset and showed strong retention. However, we do not observe offline gains between sessions. Notably, accuracy declined transiently for the novel track configurations, suggesting interference from prior training. Movement efficiency, indexed by path length, was retained and generalized to the rotated track. However, reversing movement direction impaired efficiency, revealing a movement direction effect. This effect persisted when training direction was reversed in a second study, with counterclockwise movements remaining slower and less efficient than clockwise movements. These findings show that practice produces durable and broadly transferable motor execution improvements, while inherent movement direction biases constrain how improvements generalize across contexts. New & NoteworthyThe learning principles underlying improvements in motor acuity remain less well understood than those governing other forms of motor learning. Prior work suggests that motor execution improvements show limited generalization. In contrast, the present findings demonstrate that execution-based practice can produce robust, transferable gains, while also revealing a key constraint: inherent movement direction biases that limit generalization. By characterizing learning, retention, and generalization, this work provides new insight into how motor acuity improvements compare with skill acquisition and adaptation.

neuroscience↗

bioRxiv2026

ATM safeguards DNA replication at endogenous base lesions

Poly(ADP-ribose) polymerase inhibitors (PARPi) exploit homologous recombination deficiency (HRD) in BRCA1/2-mutated cancers to induce synthetic lethality1-3. PARPi also kill cancer cells lacking the DNA damage-responsive kinase ATM4-6, however, inconsistent evidence of HRD7-9 and variable clinical responses9-11 have obscured the underlying mechanism. Here we define how PARPi induce cytotoxicity in ATM-deficient cells and reveal a critical role for ATM in regulating DNA replication. In the absence of ATM, unrestrained PRIMPOL-dependent repriming at spontaneous oxidative base adducts generates discontinuous daughter strands containing DNA gaps that activate PARP. This defect is sustained by aberrant suppression of replication fork slowing - presumably via fork reversal - by the BRCA1-A complex, whose recruitment to stalled forks is normally counteracted by ATM. The resulting gaps require homologous recombination (HR) for post-replicative repair and underlie the synthetic lethal interaction with PARPi. Suppressing repriming, reducing oxidative stress, or blocking base excision repair alleviates these defects. Collectively, our findings reveal how spontaneous base damage cooperates with replication dysfunction to drive PARPi sensitivity and establish a paradigm of post-replicative repair addiction in ATM-deficient cells. Together, our findings define a mechanistic link between oxidative DNA damage and ATM-dependent replication control, illuminating how oxidative stress may exacerbate genome instability in Ataxia Telangiectasia.

molecular biology↗

bioRxiv2026

Roots of Wheat and Rice maintain Gravitropic Setpoint Angles (GSAs)

Root growth angle is a key determinant of root system architecture, nutrient capture efficiency and therefore yield. Yet the mechanisms governing non-vertical growth in cereal roots remain poorly understood. Here, we investigated if cereal roots maintain Gravitropic Setpoint Angles (GSAs) and the hormonal regulatory processes underpinning GSA maintenance in cereals. Firstly, we found that both wheat seminal roots and rice crown roots actively return toward their original growth angles following displacement, consistent with true GSA maintenance. Next, we show that removal of a stable reference to gravity through clinorotation resulted in a characteristic outward curvature in all root types, indicating the presence of an antigravitropic offset similar to that described in Arabidopsis. Exogenous auxin treatment induced steeper rooting in both species, suggesting conserved hormonal regulatory mechanisms of GSA in both monocots and dicots. Interestingly, lateral root GSAs displayed species-specific differences: wheat laterals returned to their GSAs more effectively than rice laterals, which showed slower and incomplete responses. Together, these findings establish that cereal roots maintain GSAs through gravity-dependent and auxin-regulated mechanisms, providing a novel framework for understanding and manipulating root system architecture in monocot crops.

plant biology↗

bioRxiv2026

Large-scale Visuomotor Reaction Time Self-Testing Reveals Subtle Motor Changes in Older Adults with Subjective Cognitive Impairment

IntroductionAffordable tools for early Alzheimers disease (AD) detection could support drug development and early intervention. Subtle motor changes may indicate preclinical AD, but hand response selection and initiation speeds are understudied. This study assessed whether unsupervised, online visuomotor reaction time (RT) tests relate to subjective cognitive impairment (SCI), a validated high risk state for future conversion to AD MethodsA total of 910 participants (age 66.3{+/-}7.5, 70.8% female) completed assessments of simple and choice visuomotor RT tests at home as part of the online TAS Test protocol; they also completed the Cambridge Neuropsychological Test Automated Battery (CANTAB) episodic memory and executive function tests. Among them, 142 participants reported SCI. ResultsOn the TAS Test visuomotor tests, SCI was associated with 8.4% [1.4%, 15.4%] longer RT ( (p = .008; adjusted for task complexity), greater odds of time-out failure (OR = 1.35 [1.01, 1.81]; p = .037), and greater variance in RT (log-variance (SCI - comparison) = .094 [.028, .159]; p < .001). There were no significant differences between the SCI and comparison groups on any of the CANTAB tests. After adjusting for SCI status, none of the CANTAB tests were significantly associated with RT. DiscussionSCI was associated with longer and more variable visuomotor RT, and greater odds of time-out failure, while not being associated with tests of memory and executive function. Cognitive test scores did not explain a significant amount of variance in visuomotor RT. Taken together, these results support a hypothesis that people with SCI may be experiencing earlier visuomotor deficits that are distinct from (or precede) decline in episodic memory and executive function. Visuomotor tasks that record RT may be more sensitive to preclinical manifestations of cognitive decline than more traditional tests of cognitive function.

bioinformatics↗

bioRxiv2025

The ERCC6L2-MRI-KU complex coordinates NHEJ at staggered DNA double-strand breaks.

ERCC6L2 disease is a recessive bone marrow failure (BMF) syndrome caused by mutations in the SNF2-like putative DNA helicase ERCC6L2. While implicated in DNA replication, double strand break (DSB) repair via non-homologous end joining (NHEJ), and interstrand crosslink (ICL) repair, how ERCC6L2 supports haematopoietic longevity remains unclear. Investigating this in vivo, we find that an Ercc6l2-deficient haematopoietic stem and progenitor cell (HSPC) compartment in mice is unexpectedly resilient. Ercc6l2 loss was also tolerated in mice co-deficient for endogenous formaldehyde detoxification, which precipitates early-onset BMF in models of Fanconi anaemia. Instead, Ercc6l2-deficient mice display a mild immunodeficiency, arising from defects in immunoglobulin class-switch recombination (CSR), that synergise with shieldin-deficiency, implicating ERCC6L2 and shieldin in distinct repair mechanisms. Furthermore, we demonstrate that ERRC6L2 stimulates chromosome fusions in the context of staggered, but not blunt dysfunctional telomeres. We reconcile ERCC6L2s NHEJ function through proteomic elucidation of its endogenous interactome and AlphaFold structural modelling to reveal a complex formed of ERCC6L2 and KU that is bridged by the NHEJ accessory factor MRI/CYREN. Consequently, ERCC6L2-MRI inter-dependence characterises CSR. Together, our findings implicate the ERCC6L2-MRI complex as a KU-regulatory DNA translocase coordinating classical-NHEJ at staggered-end DSBs. We suggest that similar staggered-end breaks represent the pathological substrates driving haematopoietic failure in ERCC6L2 disease.

molecular biology↗

bioRxiv2025

Combinatorial inhibition of LSD1 and Menin induces therapeutic differentiation in AML

Acute myeloid leukemia (AML) is characterized by differentiation arrest and uncontrolled proliferation. Differentiation therapy aims to treat AML by de-repressing latent myeloid maturation programs to induce cell cycle arrest and subsequent cell death. This approach is curative in the promyelocytic AML subtype, but has met with limited success in other subtypes. Genes such as LSD1 have emerged as intriguing non-APL AML differentiation therapy targets, but results as monoagents in clinical trials have been mixed. Here, we performed differentiation-specific CRISPR screens to identify targets whose inhibition synergizes with LSD1 inhibition to induce terminal differentiation of non-APL AML cells. Intriguingly, the MLL co-factor Menin scored as the top hit. Using cell lines, primary patient samples, and mouse AML models, we find that dual inhibition of LSD1 and Menin is a highly promising approach for differentiation therapy. Mechanistically, we determine that inhibition of Menin downregulates drivers of proliferation and stemness such as MEIS1, and inhibition of LSD1 induces inflammatory and interferon-related pro-myeloid differentiation expression programs. Surprisingly, we find that this combination is effective in selected AML models without mutations in MLL or NPM1, thus nominating dual inhibition of LSD1 and Menin as an attractive therapeutic approach for a mutationally diverse set of non-APL AMLs. HighlightsO_LIInhibition of LSD1 and Menin synergizes to induce differentiation of MLL-r and MLL-WT AMLs. C_LIO_LIInhibition of Menin downregulates drivers of proliferation and stemness. C_LIO_LIInhibition of LSD1 induces differentiation-associated inflammatory and interferon responses. C_LIO_LILSD1 and Menin occupy different areas of the genome. C_LI

cancer biology↗

bioRxiv2025

Aberrant NSUN1 Activity Connects m5C RNA Modification to TDP-43 Neurotoxicity in ALS/FTD

In amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), the nuclear RNA-binding protein TDP-43 mislocalises to the cytoplasm and forms insoluble aggregates, but the mechanisms controlling this remain unclear. We define a native TDP-43 interactome in human SH-SY5Y cells and identify proteins linked to the 5-methylcytosine (m5C) RNA modification as highly enriched. Using a Drosophila model of TDP-43 pathology, we show that aberrant activity of m5C-RNA methyltransferases Nsun1 drives TDP-43-induced m5C-RNA hypermethylation, while Nsun1 downregulation alleviates TDP-43-induced degeneration, lifespan deficits, and cytoplasmic accumulation. In human cells, TDP-43 selectively interacts with NSUN1 isoform 3 independently of RNA. Furthermore, NSUN1 is nucleolar and TDP-43 is largely nucleoplasmic, yet they interact in both compartments, suggesting functional roles beyond their predominant localisations. In ALS/FTD post-mortem frontal cortex, NSUN1 isoform 3 persists, while the shorter isoform is reduced, suggesting that a pool of NSUN1 capable of contributing to pathological TDP-43 interactions remains in disease. These findings suggest that TDP-43 neurotoxicity is coupled to NSUN1 activation and m5C-RNA methylation, revealing a potential therapeutic axis in ALS/FTD. Summary blurbThe study shows that NSUN1-mediated m5C RNA methylation drives TDP-43 cytoplasmic accumulation and neurotoxicity, revealing a new pathway contributing to ALS/FTD pathogenesis.

cell biology↗

bioRxiv2025

Endothelial Adgrl2 expression and alternative splicing controls the cerebrovasculature

Central nervous system development requires parallel but interrelated processes of neural circuit assembly and vascularization. Intersecting between these two processes, the cell-adhesion G-protein coupled receptor Adgrl2 is expressed in select neuron populations where it has been found to localize and control the assembly of specific synaptic sites. Further, Adgrl2 is found to be expressed in select non-neuronal brain cells, where it is restricted to endothelial cells. Testing for Adgrl2 function in these cells, here we find that endothelial cell specific Adgrl2 deletion results in an impairment in cerebrovascular integrity. To understand how it might be possible for Adgrl2 to function independently in neuronal and endothelial contexts, we analyzed single-cell RNA sequencing datasets for differences in transcriptional identity between these cell classes. Doing so, we find that expression of Adgrl2 mRNA is subject to robust cell type-specific alternative splicing, resulting in distinct isoforms being produced in neurons compared to endothelial cells. Investigating the functional implication of this cell type-specific alternative splicing, we find that forced expression of the neuronal isoform of Adgrl2 in endothelial cells leads to alterations in cerebrovascular properties. Morphologically, endothelial cells with the non-native neuronal Adgrl2 isoform induce the formation of ectopic glutamatergic synaptic contacts onto endothelial cells, indicating alterations in the cell-cell recognition process. Functionally, in direct contrast to endothelial Adgrl2 deletion, this genetic expression switch instead enhances blood-brain barrier integrity. This overly restrictive cerebrovascular function results in dysregulation of blood to cerebrospinal fluid homeostasis, enlargement of brain ventricles, and a higher risk of hydrocephalus. As such, alternative splicing serves as a cell type specific mechanism that provides isoform specific Adgrl2 for discerning functions controlling neural circuit assembly and cerebrovascular homeostasis.

neuroscience↗

bioRxiv2025

Region-specific human brain organoids reveal synaptic and cell state drivers of glioblastoma invasion

Glioblastoma (GBM) is a highly heterogenous and malignant brain tumor, in part because it disrupts normal brain circuits to fuel its own growth and invasion. Thus, there is a need to identify the molecular features of invasive GBM cells and their regulators in the neural microenvironment. To address this in a fully human model, we engrafted patient-derived GBM cells (total n=15 independent samples) from three sources-- fresh neurosurgical resections, cell lines, and whole GBM organoids--into human induced pluripotent stem cell-derived organoids patterned to forebrain, midbrain, and spinal cord identities. GBM cells from all sources infiltrated brain organoids within 2 days post-engraftment, reaching maximal invasion by day 14. Across organoids of distinct spatial and maturational milieu, GBM cells showed a consistent reduction in astrocyte-like states and an enrichment in neuron/glia progenitor-like (NPC-like) states. These NPC-like GBM cells expressed neuronal and synaptic machinery, and tumors enriched in this transcriptomic state prior to engraftment achieved greater organoid coverage, suggesting enhanced infiltration and synaptic integration of this GBM cell type. Although GBM cell states converged across organoid types after engraftment, infiltration was greater in the forebrain than spinal cord. This is likely reflective of synaptic input from deep-layer TBR1 excitatory neurons in the forebrain, as demonstrated by a combination of rabies-based monosynaptic tracing and single-cell transcriptomics. In contrast, inhibitory neurons were the predominant synaptic partners of GBM in the spinal cord. Together, this fully human model of the neural-GBM connectome reveals how neuron-like GBM states and regionally distinct synaptic inputs cooperatively shape tumor invasion.

neuroscience↗

bioRxiv2025

beta3-adrenergic browning of pericardial adipose tissue controls cardiac function

Dysfunctional adipose tissue (AT) is strongly linked to the development of cardiovascular diseases (CVD). Accumulation of AT around vital organs is detrimental to their respective function and overall health. Although there is strong evidence linking the accumulation of pericardial AT with CVD development, a comprehensive investigation on the adaptation of pAT in obesity is scarce. Here, by applying pair-wise bottom-up proteomics in pAT of humans and mice, we found pAT presents a browning signature, as demonstrated by enrichment of mitochondria, presence of UCP1, and greater metabolic capacity compared to subcutaneous AT. In mice fed a high-fat diet or obese patients, the pAT undergoes whitening, characterized by adipocyte hypertrophy, reduced mitochondrial content, respiratory capacity, and UCP1 levels. Lipectomy of pAT from obese mice decreased pathological ventricular hypertrophy. Conversely, selective {beta}3-adrenergic agonist treatment rescued pAT browning status and is associated with improved heart structure and function, including ventricular thickness, and fibrosis in obese mice. Importantly, lipectomy of pAT abrogated the positive effects of {beta}3-adrenergic agonism in cardiac function of obese mice. Altogether, our work positions pAT as a mechanistic driver of obesity-related cardiac dysfunction and establish {beta}3-adrenergic-mediated browning of pAT as a novel therapeutic treatment strategy.

physiology↗

bioRxiv2025

Tool and hand adaptation and localization in immersive virtual reality

The human brain readily adapts movements to achieve motor goals. Most visuomotor adaptation studies use planar reaching with a cursor, where misaligned feedback leads to compensatory adjustments, reach aftereffects, and shifts in perceived hand location. Whether these effects extend to more natural settings and tool use remains unclear. In the Hand Experiment, we showed that in immersive virtual reality (VR), adaptation to 30{degrees} and 60{degrees} visuomotor rotations produced robust reach aftereffects and the expected shifts in hand localization. In the Pen Experiment, we extended this paradigm to a familiar hand-held tool, assessing localization of both the tool tip and the hand. Adaptation with the pen induced comparable or greater recalibration effects than with the hand-cursor, including shifts in both perceived tool and hand position. These findings demonstrate that visuomotor adaptation in immersive VR generalizes beyond cursor-based tasks, revealing how the sensorimotor system recalibrates internal representations of both the body and tools in realistic 3D environments. Author summary for PLOS ONEThis study shows that when people adapt their movements in virtual reality, the brain recalibrates not only the sensed position of the hand but also of familiar tools, highlighting how we update body and tool representations in everyday-like environments.

neuroscience↗

bioRxiv2025

Lead (Pb) exposure alters neural cell fate in the developing human brain

The heavy metal lead (Pb) is a developmental neurotoxicant associated with cognitive and behavioral deficits, but the cellular mechanisms underlying these impairments remain unclear. Here we show that prenatal Pb exposure biases human radial glia fate, prolonging neurogenesis and suppressing astrogenesis. We used hiPSC-derived cortical organoids, primary human fetal tissue, and in vivo xenografts to demonstrate that Pb exposure alters radial glial differentiation. Pb-exposed organoids contain a higher proportion of neurons and fewer astrocytes. We validated this differentiation bias in primary radial glia from human cortices (GW16-20), observing Pb-associated reductions in astrocyte commitment via genetic lineage tracing. This correlated with increased H3K27me3, a repressive histone modification deposited by the histone methyltransferase complex PRC2, suggesting epigenetic reprogramming as a mechanistic link between Pb and neural cell fate commitment. Our findings indicate that prenatal Pb exposure impacts lineage commitment in the developing brain, which may contribute to cognitive and behavioral impairment.

neuroscience↗

bioRxiv2025

Paired plus-minus sequencing is an ultra-high throughput and accurate method for dual strand sequencing of DNA molecules

Distinguishing real biological variation in the form of single-nucleotide variants (SNVs) from errors is a major challenge for genome sequencing technologies. This is particularly true in settings where SNVs are at low frequency such as cancer detection through liquid biopsy, or human somatic mosaicism. State-of-the-art molecular denoising approaches for DNA sequencing rely on duplex sequencing, where both strands of a single DNA molecule are sequenced to discern true variants from errors arising from single stranded DNA damage. However, such duplex approaches typically require massive over-sequencing to overcome low capture rates of duplex molecules. To address these challenges, we introduce paired plus-minus sequencing (ppmSeq) technology, in which both DNA strands are partitioned and clonally amplified on sequencing beads through emulsion PCR. In this reaction, both strands of a double-stranded DNA molecule contribute to a single sequencing read, allowing for a duplex yield that scales linearly with sequencing coverage across a wide range of inputs (1.8-98 ng). We benchmarked ppmSeq against current duplex sequencing technologies, demonstrating superior duplex recovery with ppmSeq, with a rate of 44%{+/-}5.5% (compared to [~]5-11% for leading duplex technologies). Using both genomic as well as cell-free DNA, we established error rates for ppmSeq, which had residual SNV detection error rates as low as 7.98x10-8 for gDNA (using an end-repair protocol with dideoxy nucleotides) and 3.5x10-7{+/-}7.5x10-8 for cell-free DNA. To test the capabilities of ppmSeq for error-corrected whole-genome sequencing (WGS) for clinical application, we assessed circulating tumor DNA (ctDNA) detection for disease monitoring in cancer patients. We demonstrated that ppmSeq enables powerful tumor-informed ctDNA detection at concentrations of 10-4 across most cancers, parts per million sensitivity in cancers with high mutation burden, and further increased sensitivity with higher sequencing depth. We then leveraged genome-wide trinucleotide mutation patterns characteristic of urothelial (APOBEC3-related and platinum exposure-related signatures) and lung (tobacco-exposure-related signatures) cancers to perform tumor-naive ctDNA detection, showing that ppmSeq can identify a disease-specific signal in plasma cell-free DNA without a matched tumor, and that this signal correlates with imaging-based disease metrics. Altogether, ppmSeq provides an error-corrected, cost-efficient and scalable approach for high-fidelity WGS that can be harnessed for challenging clinical applications and emerging frontiers in human somatic genetics where high accuracy is required for mutation identification.

genomics↗

bioRxiv2025

Volumetric Differences of Thalamic Nuclei are Associated with Post-Trauma Psychopathology

Previous investigations of whole thalamus and thalamic nuclei volumes in post-trauma psychopathology have been sparse, limited in scope, and yielded inconsistent results. To address this, volumetric estimates of whole thalamus and thalamic nuclei were obtained from structural brain MRI scans from 2,058 participants across 20 worldwide sites in the ENIGMA PTSD working group. Thalamic volumes were compared between trauma-exposed participants with posttraumatic stress disorder (PTSD) (n=238), major depressive disorder (MDD) (n=184), comorbid PTSD+MDD (n=618), and trauma-exposed control participants (n=1,018). PTSD and MDD symptom severity, PTSD symptom clusters, and childhood trauma were similarly examined for associations with thalamic volume. Participants with PTSD only compared to controls had smaller thalamic nuclei volumes in sensorimotor nuclei, including the parafascicular (Pf), ventral anterior magnocellular (VAmc), medial pulvinar (PuM), and anterior pulvinar (PuA) nuclei of the thalamus. MDD only and comorbid PTSD+MDD participants exhibited smaller mediodorsal thalamus volumes compared to controls. Overall PTSD and MDD symptom severity negatively correlated with the volume of the mediodorsal thalamus. A significant interaction between PTSD and MDD severity was found, such that MDD severity was positively associated with thalamic volume only among individuals with high PTSD severity. Avoidance and hyperarousal symptoms of PTSD were positively associated with thalamic volume, while re- experiencing and negative mood/cognition symptoms were negatively associated with thalamic volume. Childhood physical and emotional abuse were positively and negatively associated with thalamic volume, respectively. Whole thalamus volume and volumes of the sensorimotor and limbic thalamus may play an important role in the development of PTSD and MDD in the aftermath of trauma exposure. The interaction between PTSD and MDD symptoms and contrasting effects across PTSD symptom clusters and types of childhood adversity suggests multiple neurobiological mechanisms are involved in shaping thalamic volume post-trauma.

neuroscience↗

bioRxiv2025

Pan-Cancer PDOs Preserve Tumor Heterogeneity and Uncover Therapeutic Vulnerabilities

We developed a tumor-matched, pan-cancer patient-derived organoid (PDO) platform comprising 220 PDOs from 190 patients across 15 cancer types to advance functional precision oncology. Our comprehensively characterized PDOs showed 93% histopathology concordance, 80% median genomic concordance for driver mutations, and a 0.85 median gene expression correlation with parent tumors. Gene expression in PDOs remained stable across [≥] 10 passages, supporting reproducibility for long-term drug screening. Even PDOs with low genomic concordance retained oncogenic drivers, supporting their use as disease models. Clonality analysis revealed that 85% of PDOs preserved dominant tumor clones. Higher genomic concordance was associated with greater clonal similarity, while lower genomic concordance was associated with clonal divergence. Functional assays showed that 58% of PDOs from a subset of patients ineligible for FDA-approved PARP inhibitors responded to Talazoparib, with sensitivity linked to alterations in DNA damage repair. Combination screens revealed drugs that effectively overcame resistance, especially in TP53-mutant PDOs. In summary, our platform supports investigation of targeted therapies, identification of molecular features linked to drug sensitivity, and translational discovery, offering insights into personalized cancer treatment beyond current biomarker guidelines.

cancer biology↗