Biology subjects

Wong, E.

Publications and source records attributed to Wong, E..

At least 19 recordsLinked to original sources

bioRxiv2026

Genetic dissection of DNA damage response pathways reveals divergent modulation of PD-1 blockade response in colorectal cancer

Although MSI-H/dMMR colorectal cancers and the rarer subset with pathogenic POLE exonuclease domain mutations can benefit from immune checkpoint blockade, MSS colorectal cancer remains largely refractory, raising the question of whether other DNA damage response defects may also confer sensitivity to immunotherapy. Here, we performed a systematic in vivo screen of recurrently altered DNA damage response (DDR) genes to determine whether distinct repair defects modulate response to PD-1 blockade in MSS colorectal cancer. Using a genome-edited murine model in immunocompetent mice, we found that DDR perturbations produced heterogeneous outcomes, indicating that the immunologic consequences of DNA repair deficiency may be pathway-specific. Loss of Brca1, Brca2, Atm, and Fanca enhanced response to anti-PD-1 therapy, whereas Atr and Msh6 deficiency were associated with poor benefit. Brca1/2-deficient tumors showed the strongest regressions, supporting a link between core homologous recombination deficiency and increased immune sensitivity. In an independent cohort of colorectal cancer patients treated with ICB from the MSK-IMPACT Clinical Sequencing Cohort, BRCA1 alterations were associated with improved survival status, while FANCA alterations showed a similar trend. Because ATM is therapeutically targetable, we evaluated pharmacologic ATM inhibition in MSS colorectal cancer. Sustained ATM inhibition induced immune-related transcriptional programs in vitro and, when administered as a pre-treatment in vivo, enhanced the antitumor activity of PD-1 blockade. Together, these findings suggest that selected DDR alterations may represent candidate biomarkers of immunotherapy sensitivity, while ATM inhibition may provide a rational combination strategy to enhance PD-1 blockade in MSS colorectal cancer.

cancer biology↗

bioRxiv2026

Decoupled evolution of antimicrobial repertoires in lichen-forming fungi

Lichens are stable multipartite symbioses in which a fungal mycobiont associates with one or more photosynthetic partners while hosting complex microbiomes. Secreted antimicrobial proteins (AMPs) have recently emerged as important mediators of fungal interactions with surrounding microbial communities, yet their evolutionary distribution and diversity in lichen-forming fungi remain largely unexplored. Here, we predicted AMP repertoires across 80 phylogenetically diverse lichen-forming fungi representing three photobiont association types. Contrary to our expectation, average repertoire size did not differ between chlorolichens, cyanolichens and tripartite lichens (363 AMPs per species overall), even though repertoire size varied almost five-fold among species and 23 AMP families showed photobiont-associated enrichment or depletion. This quantitative conservation masks extensive compositional turnover, with families occurring in at least 90% of species coexisting alongside numerous lineage-specific groups. Pooled population sequencing of 11 Umbilicaria phaea populations along two elevational gradients further revealed recurrent AMP presence-absence variation within a single species. In contrast to AMPs, biosynthetic gene clusters (BGCs) and carbohydrate-active enzymes (CAZymes), which include known antimicrobials, were markedly reduced in cyanolichens relative to chlorolichens. Lichen-forming fungi thus combine a conserved baseline antimicrobial protein capacity, whose composition nonetheless turns over between lineages and populations, with chemical and enzymatic systems that differentiate along photobiont association and may support ecological and lifestyle-associated adaptation.

microbiology↗

bioRxiv2026

LifeSciBench: Evaluating Language Models on Realistic, Expert-Level Tasks in the Life Sciences

We introduce LifeSciBench, a benchmark of 750 expert-authored tasks designed to evaluate whether language models can handle realistic life science research work. The majority of existing life sciences benchmarks have a narrow scope or are purely knowledge-based, and therefore fail to capture the complexity of real-world research, which often involves ambiguities and requires the accurate execution of multiple dependent judgment calls. Additionally, almost all existing benchmarks span at best a small collection of subdomains within the life sciences; there is at present no existing life sciences benchmark with both the requisite breadth and depth required to convincingly measure proficiency in real-world professional research settings. LifeSciBench addresses this gap by spanning seven representative scientific workflows and seven life science domains, with each constituent task paired with a human expert-written rubric. Across five frontier and domain-specialized models, GPT-Rosalind performs best, with a task-weighted mean normalized rubric score of 0.576 and a task-weighted response pass rate of 36.1% (response-level values are first averaged within each task, and the resulting task-level values are then averaged with equal weight). LifeSciBench remains unsaturated, with 171 tasks (22.8%) having no observed passing response from any evaluated model and 261 tasks (34.8%) having a best-model pass rate below 20%. LifeSciBench therefore serves as a high-resolution evaluation of practical scientific reasoning and operational decision-making in the life sciences.

bioinformatics↗

bioRxiv2026

Mitochondrial competence determines responses to metabolic interventions during aging

Cellular responses to metabolic interventions vary across physiological contexts, but the basis for this variability remains unclear. Here, we show that mitochondrial competence determines whether cells can engage adaptive metabolic states that support survival during aging. Using 4-methylbenzoic acid (4-MBA), identified as a lifespan-extending compound, as a perturbation probe, we find that modulation of Target of Rapamycin Complex 1 (TORC1) signaling induces a shift from anabolic growth to maintenance-associated metabolism; however, signaling output alone does not predict outcomes. Instead, survival closely correlates with mitochondrial function, with its disruption abolishing adaptive responses. Genetic and biochemical analyses define a mitochondrial regulatory circuit that constrains signaling dynamics and governs state transitions. This regulatory logic is conserved across mammalian systems and operates under oxidative stress, replicative aging, and Hutchinson-Gilford progeria syndrome (HGPS), extending to proliferative cancer cells. These findings establish mitochondrial competence as a key determinant of cellular responsiveness and provide a framework for understanding context-dependent effects of metabolic interventions during aging.

cell biology↗

bioRxiv2026

Principles of in situ protein sequencing: expansion microscopy-adapted Edman degradation and amino acid recognition

The ability to map protein identity, with resolution sufficient to infer interactions, would support analysis of how proteins work together, or malfunction, in biological processes and diseases. Although several emerging technologies aim towards single-molecule protein sequencing, they require proteins to be removed from the nanoscale spatial context of cells and tissues. Expansion microscopy (ExM) has facilitated a diversity of chemical analyses by isotropically separating molecules throughout a specimen after permeation via a charged hydrogel, followed by gel swelling. Here, we adapt key protein sequencing steps - Edman degradation and amino acid recognition - to the ExM gel context. Using testbed peptides in ExM gels, we show that N-terminal amino acids can be recognized over multiple cycles of in-gel Edman degradation. This principle-oriented study demonstrates sequencing chemistry on defined synthetic constructs, rather than endogenous proteins in biological samples. These results establish principles of in situ protein sequencing and provide a framework for future in situ protein sequencing developments, including the development of higher specificity and affinity amino acid binders.

biochemistry↗

bioRxiv2026

Proteo-transcriptomics and morphometrics of teleost cardiac cells define regulatory networks and exercise-induced cardiomyocyte hypertrophy and hyperplasia

Zebrafish and medaka are powerful cardiovascular models, yet cellular and molecular investigations of adult heart cells have been constrained by suboptimal dissociation and characterization methods. To overcome these barriers, we developed a physiological-temperature workflow that generates high-yield, viable single-cell suspensions for FACS, imaging, and low-input molecular profiling. Using transgenic fluorescent reporters, we consistently isolate [~]6,000 cardiomyocytes per adult zebrafish ventricle and [~]12,000 from medaka, preserving cellular, structural, and molecular integrity. Single-cell morphometrics revealed cardiomyocyte heterogeneity and demonstrated that swimming exercise induces both hypertrophy and hyperplasia, while ventricular injury triggers expansion of regenerative gata4 cardiomyocytes. We integrated proteomics and RNA-seq from FACS-purified cells to construct cell-type-specific proteo-transcriptomic atlases. Functional enrichment, transcription factor, and network analyses identified protein hubs and regulatory circuits defining cardiomyocyte and endothelial cell identity. Our platform delivers cell-type-resolved molecular datasets of adult teleost cardiac cells, establishing a systems-level resource for heart regeneration research, cardiovascular disease modeling, and drug discovery. TeaserA robust workflow for isolation, FACS, imaging, and multi-omics profiling of cardiomyocytes and cardiac endothelial cells in fish. HIGHLIGHTSO_LITemperature-optimized dissociation and standardized FACS enable high-yield isolation of viable cardiac cells. C_LIO_LISingle-cell imaging uncovers morphological heterogeneity in adult ventricular cardiomyocytes. C_LIO_LISustained exercise induces both hypertrophy and hyperplasia of zebrafish cardiomyocytes. C_LIO_LIProteo-transcriptomics defines core molecular programs and interaction networks in cardiomyocytes and endothelial cells. C_LI

systems biology↗

bioRxiv2025

PLD1-Dependent Regulation of Synaptic Integrity: Implications for Cognitive Resilience and Alzheimer's Disease Pathogenesis

Dysregulation of Phospholipase D1 (PLD1) has been implicated in the progression of neurodegenerative diseases, including Alzheimers Disease (AD). This study investigated PLD1 signaling in synaptic integrity and cognition during aging and in a late-onset AD mouse model, hypothesizing differential effects of PLD1 modulation on synaptic vulnerability. AAV2-mediated gene transfer was employed to overexpress (PLD1 OXP) or attenuate (PLD1 ATT) PLD1 in aged wild-type (WT) and 3xTg-AD mice. To validate these constructs, differentiated PC12 cells were utilized. Within these cells, a model of post-mitotic neurons, PLD1 OXP exhibited a notable reduction in both the average neurite length and the percentage of neurite-bearing cells. This suggests that elevated PLD1 activity exerts a significant influence on neurite outgrowth. Conversely, PLD1 ATT did not inhibit neurite formation, indicating it is not detrimental at the cellular level. These cellular findings paralleled in vivo observations - electrophysiological studies revealed PLD1 OXP impaired long-term potentiation (LTP) and synaptic transmission, particularly in aged WT mice, whereas PLD1 ATT improved synaptic function in 3xTg-AD mice. Behaviorally, PLD1 ATT enhanced spatial working memory and reduced anxiety-like behavior, notably in 3xTg-AD mice. These results highlight that tight PLD1 regulation is vital for maintaining synaptic integrity and cognitive resilience. Thus, PLD1 attenuation may serve as an important complement to immunotherapeutic approaches by strengthening synaptic resilience in neurodysfunctional states, including AD and related dementia (ADRD).

neuroscience↗

bioRxiv2025

MDFIC2 is a sensory neuron-specific PIEZO channel auxiliary subunit

PIEZO channels are critical for sensory mechanotransduction. While MyoD-family inhibitor proteins were identified as PIEZO1 auxiliary subunits, their broader regulatory roles, particularly in sensory cells, remained unclear. Here we demonstrate native MDFIC and MDFI regulate endogenous PIEZO channel currents in various non-sensory cell types. However, neither MDFIC nor MDFI are expressed in primary sensory neurons. In these cell types we identified an uncharacterised member of this family, Mdfic2/Gm765, that shares the ability to physically bind to PIEZO1 and PIEZO2. MDFIC2 is selectively expressed in subsets of mechanosensitive neurons, including dorsal root ganglia, trigeminal ganglia, and vagal sensory neurons. Like its paralogues, MDFIC2 alters PIEZO1/2 mechanosensitivity and inactivation kinetics, converting them into high-threshold slowly inactivating mechanoreceptors. Extensive cryo-EM reveals a conserved binding pocket for these auxiliary subunits in the pore modules of both PIEZO1 and PIEZO2 mediated by the post-translationally modified distal C-termini of MyoD-family inhibitor proteins. This provides a comprehensive structural and functional characterisation of MyoD-family inhibitor proteins as PIEZO1/2 channel regulators and offers new insights into sensory physiology and mechanical pain mechanisms.

biophysics↗

bioRxiv2025

Comprehensive public health surveillance identifies novel human cryptosporidiosis subtypes and genetic diversity associated with swimming pool outbreaks in Australia, Victoria (2018-2024)

Cryptosporidium and Giardia are major causes of gastrointestinal illness globally. In Australia, cryptosporidiosis is a nationally notifiable disease, yet molecular characterisation of clinical cases is rarely performed, limiting the capacity to identify outbreaks, trace sources and assess zoonotic risk. During 2024 there was a 273% cases increase in Australia, the third country with the highest increase. We present the first comprehensive molecular investigation of human Cryptosporidium and Giardia infections in the state of Victoria, Australia. We analysed faecal samples collected between 2018 and 2024. Positive samples were subtyped and parasite load was estimated. Of the 2,330 samples tested, 225 were positive for Cryptosporidium and nine for Giardia. Seven Cryptosporidium species and 24 subtypes were identified, including multiple novel or regionally unique subtypes. C. hominis was the predominant species (85%), and three subtypes associated with 11 recreational water outbreaks in 2024. Based on spatiotemporal overlap and subtypes, 52 cases were inferred to represent undetected outbreak-associated infections. Several C. parvum subtypes reflected probable zoonotic transmission, two subtypes were associated with a childcare and camp outbreak. Six C. hominis subtypes and eight subtypes overall were reported for the first time in Australia. Globally novel subtypes of C. occultus, C. fayeri and C. meleagridis in human hosts is reported. The study reveals high diversity of Cryptosporidium subtypes in human infections in Victoria. This study demonstrates the potential for molecular surveillance to inform public health interventions when integrated with epidemiological data. These findings support One Health approaches to outbreak detection and source attribution. IMPORTANCECryptosporidium is a nationally notifiable pathogen in Australia, yet routine genotyping is not performed, limiting outbreak detection and source attribution. This study represents the first large-scale molecular surveillance of human Cryptosporidium and Giardia infections in southern Australia. By integrating species- and subtype-level typing with epidemiological metadata, we demonstrate the substantial diversity of infective lineages, including several novel or zoonotic genotypes. Our findings highlight the critical role of molecular tools in tracking transmission pathways, supporting outbreak investigation and informing public health responses. These data provide a foundation for incorporating routine genotyping into national surveillance strategies for parasitic enteropathogens. This study presents the first epidemiological study of the 2024 outbreaks in Australia. The global comparison of cases during this period also highlights potential large-scale disease dynamic of public health importance.

molecular biology↗

bioRxiv2025

A Complete Telomere-to-Telomere Diploid Reference Genome for Indian Population

Human reference genomes have been instrumental in advancing genomic and biomedical research, but South and Southeast Asian populations are underrepresented, despite accounting for a large proportion of world population. As a part of effort on generating reference genomes for these populations, we present the first gapless, telomere-to-telomere (T2T) diploid genome assembly created by using a trio sample set of Indian ancestry (I002C), with NG50 of 154.89 Mb and 146.27 Mb for the maternal and paternal haplotypes, including the fully assembled rDNA array for the maternal chromosome 21 and Y chromosome. With the Merqury QVs of 82.05, 83.08 and 82.64 for the maternal, paternal and haploid assemblies respectively, I002C represents the highest-quality human genome assembled in both diploid and haploid forms to date. Compared to CHM13, the I002C genome displays substantial sequence diversity, resulting in 14,943 structural variants, including 3,236 novel variants absent from public databases. Analysis of trio-phased haplotypes further revealed elevated inter-haplotype divergence within centromeric and subtelomeric regions, along with identification of differentially methylated regions (DMRs) as candidates for novel imprinting loci. As a result of substantial SVs between them, I002C is a more suitable reference than CHM13 for the genomic analysis of South Asian samples with less reference bias and better performance in mapping and variant calling, particularly for long read sequencing data. As the first high-quality T2T diploid reference genome for Indian, the largest worlds population, I002C contributes to the growing set of population-specific reference genomes and helps to overcome a significant gap in human genome diversity.

genomics↗

bioRxiv2025

Modulating populational variance of methyl-guanine methyl transferase expression through miR-181d degradation: a novel mechanism of temozolomide resistance

Intratumoral heterogeneity plays a pivotal role in cancer evolution, providing the substrate for adaptation to selective pressures, including treatment with chemotherapy. Here, we show that micro-RNA regulation of variance in the expression of the DNA repair protein methyl-guanine methyl transferase (MGMT) contributes to this heterogeneity and acquired therapeutic resistance. In cell lines derived from glioblastomas, the most common form of primary brain tumor, treatment with standard-of-care temozolomide chemotherapy triggers a feed-forward loop between polyribonucleotide nucleotidyltransferase 1 (PNPT1) and miR-181d, an MGMT regulating miRNA, expediting miR-181d degradation. This degradation requires the activation of Ataxia Telangiectasia and Rad3-related (ATR) kinase. The degradation of miR-181d in glioblastoma cells increased both the mean and the variance of MGMT expression in the cell population. Subclone reconstituted cell populations with similar populational mean MGMT levels but with differences in the variance of MGMT expression exhibited differential temozolomide sensitivity, with the higher MGMT variance population showing increased resistance. This resistance is suppressed by exogenously transfected miR-181d. These findings suggest a key role for miRNA in regulating intra-tumoral heterogeneity through modulation of key DNA repair enzymes and provide a compelling rationale for miRNA delivery as a platform for glioblastoma therapy. Significance StatementThis study demonstrates a mechanistic link between a feed-forward loop mediating microRNA degradation and cell-to-cell variance in gene expression, and the contribution of this mechanism to intratumoral heterogeneity and therapeutic resistance. We show that when glioblastoma, the most common form of adult primary brain tumor, is treated with standard-of-care chemotherapy, temozolomide, a feed-forward loop between miR-181d and PNPT1 is initiated, causing rapid degradation of miR-181d. This degradation increases the cell-to-cell variability in methyl-guanine methyl transferase (MGMT) expression, expanding intra-tumoral heterogeneity and contributing to acquired temozolomide resistance. This process can be suppressed by therapeutic delivery of microRNA, providing compelling considerations for clinical translation.

cancer biology↗

bioRxiv2025

National Scale Genomic Engine for Precision Medicine: Singapore PRECISE-SG100K Experience

The whole-of-nation National Precision Medicine (NPM) programme in Singapore aims to integrate a genomics backbone into standard healthcare. Phase II recently completed the sequencing of 102,202 genomes from the diverse Asian population of Singapore, generating the PRECISE-SG100K dataset. The scaling from the pilot SG10K_Health phase to national production scale demanded a central shift: from earlier generation workflows to an integrated, industrial grade "Genomic Engine". The initiative was co-developed under a public-private partnership model. Our Genomic Engine was designed with an automation-first approach, as a modular platform linking a purpose-built, centralized biospecimen processing and DNA extraction facility (EXTRACT Lab), a high-throughput sequencing operation (Illumina Data Generation Infrastructure, IDGI), with the cloud-native analytics (GRIDS) running on a fully integrated PRECISE Sample Tracking System (STS). Here we describe the key design features of our Genomic Engine. Highlighting the Stress Test phase where the automated DNA extraction yields failed to meet expectations. We share the operational adjustments which were implemented to stabilize production and resulted in a three-fold reduction in sample triage due to improved DNA recovery. Without establishing the operational guardrails during the Stress Test, the Engine would not have reached a steady-state production evidenced by >3,000 genomes per month, median turnaround time 11 days, 99.6% operational success, within in six months. This framework offers a practical blueprint for other population-scale initiatives seeking to bridge the gap between scientific piloting and industrial delivery.

genomics↗

bioRxiv2025

The KRAB-Zinc Finger protein ZKSCAN3 represses enhancers via embedded retrotransposons

Gene cis-regulatory sequences are increasingly recognised as containing "domesticated" transposable elements that impact their function. The KRAB Zinc Finger Protein (KZFP) family of transcription factors is typically associated with transposable element silencing through establishment of heterochromatin. Here, using acute protein depletion in embryonic stem cells, we reveal that the KZFP ZKSCAN3 represses enhancer activity through targeting enhancer-embedded retrotransposons and that ZKSCAN3-mediated repression does not rely on the induction of heterochromatin. ZKSCAN3, which exhibits strong genetic association with the neurodevelopmental disorder schizophrenia, operates during neural differentiation and is necessary for proper cell specification and expression of genes that regulate axon guidance, neuronal motility and pathfinding. These findings define ZKSCAN3 as an enhancer regulator and uncover a heterochromatin-independent KZFP. Additionally, they exemplify how a KZFP epigenetically regulates enhancers in a native setting and highlight how transposable elements and their KZFP binders have shaped gene expression networks.

molecular biology↗

bioRxiv2024

Epigenetic restoration of differentiation competency via reversal of epiblast regionalisation

Although the epiblast in the embryo has the capacity to generate all tissues of the body, its in vitro counterparts often exhibit differentiation biases, posing significant challenges for both basic research and translational applications involving pluripotent stem cells (PSCs). The origins of these biases remain incompletely understood. In this study, we identify PSC differentiation biases as arising from fluctuations in repressive and activating histone posttranslational modifications, leading to the acquisition of a caudal epiblast-like phenotype. We present a novel approach to overcome this bias using a chemical chromatin restoration (CHR) treatment. This method restores transcriptional programs, chromatin accessibility, histone modification profiles, and differentiation potential, effectively recapitulating the competent anterior epiblast-like state. Furthermore, we propose that a high bivalency state is a defining feature of the anterior human epiblast. We suggest that fluctuations in histone modification marks drive epiblast regionalization, ultimately shaping cellular responses to differentiation cues.

developmental biology↗

bioRxiv2024

Mapping DNA Methylation to Cardiac Pathologies Induced by Beta-Adrenergic Stimulation in a Large Panel of Mice

BackgroundHeart failure (HF) is a leading cause of morbidity and mortality worldwide, with over 18 million deaths annually. Despite extensive research, genetic and environmental factors contributing to HF remain complex and poorly understood. Recent studies suggest that epigenetic modifications, such as DNA methylation, may play a crucial role in regulating HF-associated phenotypes. In this study, we leverage the Hybrid Mouse Diversity Panel (HMDP), a cohort of over 100 inbred mouse strains, to investigate the role of DNA methylation in HF progression. ObjectiveWe aim to identify epigenetic modifications associated with HF by integrating DNA methylation data with gene expression and phenotypic traits. Using isoproterenol (ISO)-induced cardiac hypertrophy and failure in HMDP mice, we explore the relationship between methylation patterns and HF susceptibility. MethodsWe performed reduced representational bisulfite sequencing (RRBS) to capture DNA methylation at single-nucleotide resolution in the left ventricles of 90 HMDP mouse strains under both control and ISO-treated conditions. We identified differentially methylated regions (DMRs) and performed an epigenome-wide association study (EWAS) using the MACAU algorithm. We identified likely candidate genes within each locus through integration of our results with previously reported sequence variation, gene expression, and HF-related phenotypes. In vitro approaches were employed to validate key findings, including gene knockdown experiments in neonatal rat ventricular myocytes (NRVMs). We also examined the effects of preventing DNA methyltransferase activity on HF progression. ResultsOur EWAS identified 56 CpG loci significantly associated with HF phenotypes, including 18 loci where baseline DNA methylation predicted post-ISO HF progression. Key candidate genes, such as Prkag2, Anks1, and Mospd3, were identified based on their epigenetic regulation and association with HF traits. In vitro follow-up on a number of genes confirmed that knockdown of Anks1 and Mospd3 in NRVMs resulted in significant alterations in cell size and blunting of ISO-induced hypertrophy, demonstrating their functional relevance in HF pathology. Furthermore, treatment with the DNA methyltransferase inhibitor RG108 in ISO-treated BTBRT mice significantly reduced cardiac hypertrophy and preserved ejection fraction compared to mice only treated with ISO, highlighting the therapeutic potential of targeting DNA methylation in HF. Differential expression analysis revealed that RG108 treatment restored the expression of several methylation-sensitive genes, further supporting the role of epigenetic regulation in HF. ConclusionOur study demonstrates a clear interplay between DNA methylation, gene expression, and HF-associated phenotypes. We identified several novel epigenetic loci and candidate genes that contribute to HF progression, offering new insights into the molecular mechanisms of HF. These findings underscore the importance of epigenetic regulation in cardiac disease and suggest potential therapeutic strategies for modifying HF outcomes through targeting DNA methylation.

genomics↗

bioRxiv2024

ATP-dependent remodeling of chromatin condensates uncovers distinct mesoscale effects of two remodelers

ATP-dependent chromatin remodeling enzymes mobilize nucleosomes, but how such mobilization affects chromatin condensation is unclear. Here, we investigate effects of two major remodelers, ACF and RSC using chromatin condensates and single-molecule footprinting. We find that both remodelers inhibit the formation of condensed chromatin. However, the remodelers have distinct effects on pre-formed chromatin condensates. ACF spaces nucleosomes without de-condensing the chromatin, explaining how ACF maintains nucleosome organization in transcriptionally repressed genomic regions. In contrast, RSC catalyzes ATP-dependent de-condensation of chromatin. Surprisingly, RSC also drives micron-scale movements of entire condensates. These newly uncovered activities of RSC explain its central role in transcriptional activation. The biological importance of remodelers may thus reflect both their effects on nucleosome mobilization and the corresponding consequences on chromatin dynamics at the mesoscale.

biochemistry↗

bioRxiv2024

Aging limits stemness and tumorigenesis in the lung by reprogramming iron homeostasis

Aging is associated with a decline in the number and fitness of adult stem cells1-4. Aging-associated loss of stemness is posited to suppress tumorigenesis5,6, but this hypothesis has not been tested in vivo. Here, using physiologically aged autochthonous genetically engineered mouse models and primary cells7,8, we demonstrate aging suppresses lung cancer initiation and progression by degrading stemness of the alveolar cell of origin. This phenotype is underpinned by aging-associated induction of the transcription factor NUPR1 and its downstream target lipocalin-2 in the cell of origin in mice and humans, leading to a functional iron insufficiency in the aged cells. Genetic inactivation of the NUPR1--lipocalin-2 axis or iron supplementation rescue stemness and promote tumorigenic potential of aged alveolar cells. Conversely, targeting the NUPR1-- lipocalin-2 axis is detrimental to young alveolar cells via induction of ferroptosis. We find that aging-associated DNA hypomethylation at specific enhancer sites associates with elevated NUPR1 expression, which is recapitulated in young alveolar cells by inhibition of DNA methylation. We uncover that aging drives a functional iron insufficiency, which leads to loss of stemness and tumorigenesis, but promotes resistance to ferroptosis. These findings have significant implications for the therapeutic modulation of cellular iron homeostasis in regenerative medicine and in cancer prevention. Furthermore, our findings are consistent with a model whereby most human cancers initiate in young individuals, revealing a critical window for such cancer prevention efforts.

cancer biology↗

bioRxiv2024

Next generation AMA1-based plasmids for enhanced heterologous expression in filamentous fungi

Episomal AMA1-based plasmids are increasingly used for expressing biosynthetic pathways and CRISPR/Cas systems in filamentous fungi cell factories due to their high transformation efficiency and multicopy nature. However, gene expression from AMA1 plasmids has been observed to be highly heterogeneous in growing mycelia. To overcome this limitation, here we developed next-generation AMA1-based plasmids that ensure homogenous and strong expression. We achieved this by evaluating various degradation tags fused to the auxotrophic marker gene on the AMA1 plasmid, which introduces a more stringent selection pressure throughout multicellular fungal growth. With these improved plasmids, we observed in Aspergillus nidulans a 5-fold increase in the expression of a fluorescent reporter, a doubling in the efficiency of a CRISPRa system for genome mining, and a up to a 10-fold increase in the production of heterologous natural product metabolites. This strategy has the potential to be applied to diverse filamentous fungi.

synthetic biology↗