Biology subjects

Lane, S.

Publications and source records attributed to Lane, S..

12 recordsLinked to original sources

bioRxiv2026

Dnmt3a-mutant Leukemia Stem Cells evade chemotherapy through enforced quiescence in Npm1c-Flt3ITD Acute Myeloid Leukemia

Concurrent mutations in DNMT3A, NPM1, and FLT3 define a high-risk subtype of acute myeloid leukemia (AML) associated with increased relapse risk and inferior survival following standard chemotherapy. However, the mechanisms by which DNMT3A mutations promote treatment resistance in NPM1c-FLT3ITD AML remain unclear. Using genetically engineered murine models of Npm1c-Flt3ITD AML with or without Dnmt3aR878H (homologous to human DNMT3AR882H), we demonstrate that Dnmt3aR878H promotes chemotherapy resistance through epigenetic regulation of leukemia stem cell (LSC) quiescence. Integrated transcriptomic and epigenetic profiling revealed coordinated remodeling of DNA methylation and chromatin accessibility in LSC-enriched populations, characterized by preferential hypomethylation and increased accessibility at loci associated with stemness and quiescence programs. These data were confirmed in human DNMT3AR882H-NPM1c-FLT3ITD AML datasets with enrichment of quiescence-associated and stem cell enriched transcriptional programs. Conversely, Dnmt3a-mutant LSCs retained sensitivity to the cell-cycle independent regimen venetoclax plus azacitidine, but residual LSCs exhibited transcriptional plasticity and reversion to a de-differentiated state. We have identified LSC heterogeneity spanning primitive hematopoietic stem cell (HSC)-and progenitor-like states and our data demonstrate preferential maintenance of a quiescent HSC-like LSC subpopulation in Dnmt3aR878H-mutant AML following chemotherapy treatment. Pharmacologic induction of cell-cycle entry using pegylated interferon (pegIFN) disrupted the quiescent LSC state and restored chemotherapy sensitivity, identifying quiescence as a reversible and therapeutically actionable mechanism of resistance. These findings identify DNMT3A-mediated epigenetic regulation of LSC quiescence as a conserved mechanism of standard chemotherapy resistance and position therapeutic reactivation of quiescent LSCs as a promising strategy to overcome chemotherapy resistance and improve outcomes in high-risk DNMT3A-mutant AML.

cancer biology↗

bioRxiv2026

Spatial atlas of the ovary identifies molecular events in primordial follicle activation in humans

The human ovarian reserve is established prenatally, when granulosa cells encapsulate germ cells to form a pool of quiescent primordial follicles, that ultimately determines the female reproductive and endocrine lifespan. From birth to menopause, subsets of these quiescent follicles, located in the thin outer ovarian cortex, are activated and undergo a growth programme with progressive inward migration to the inner cortex, before either undergoing atresia or, during reproductive age, ovulation. Disruption of this process may lead to infertility, metabolic disorders and early menopause, yet early follicle development remains largely poorly understood. Here, we generate the most comprehensive single-cell and spatial multiomics atlas of the human pre- and postnatal ovarian cortex, integrating transcriptomic and chromatin profiles from over four million cells obtained from fetal and newly profiled pediatric and adult donors. We resolve the early granulosa cell trajectory at unprecedented resolution and identify a retinoic acid-associated regulatory switch accompanying follicle activation. We further show that stromal fibroblasts are not homogeneous, but instead form a dynamic scaffold establishing previously unrecognised morphogen and paracrine gradients that organise the cortex into functional niches supporting quiescent, growing, and atretic follicles. Finally, we identify ovarian lipid associated macrophages (oLAMs) that localise around follicles and are likely to support tissue remodelling during folliculogenesis. Together, this atlas provides a foundational blueprint for human ovarian development and homeostasis, and provides a framework for improving strategies in fertility preservation and in vitro follicle maturation.

cell biology↗

bioRxiv2025

Targeted control of gene expression using CRISPR-associated endoribonucleases

CRISPR-associated endoribonucleases (Cas RNases) cleave single-stranded RNA in a highly sequence-specific manner, by recognizing and binding to short RNA sequences known as direct repeats (DRs). Here we investigate the potential of exploiting Cas RNases for the regulation of target genes with one or more DRs introduced into the 3 untranslated region, an approach we refer to as DREDGE (direct repeat-enabled down-regulation of gene expression). The DNase-dead version of Cas12a (dCas12a) was identified as the most efficient among 5 different Cas RNases tested and was subsequently evaluated in doxycycline-regulatable systems targeting either stably expressed fluorescent proteins or an endogenous gene. DREDGE performed superbly in stable cell lines, resulting in up to 90% downregulation with rapid onset, notably, in a fully reversible manner. Successful control of an endogenous gene with DREDGE was demonstrated in two formats, including one wherein both the DR and the transgene driving expression of dCas12a were introduced in one step by CRISPR-Cas. Our results establish DREDGE as an effective method for regulating gene expression in a targeted, highly selective, and fully reversible manner, with several advantages over existing technologies.

molecular biology↗

bioRxiv2025

A Generalized Platform for Artificial Intelligence-powered Autonomous Protein Engineering

Proteins are the molecular machines of life with numerous applications in energy, health, and sustainability. However, engineering proteins with desired functions for practical applications remains slow, expensive, and specialist-dependent1-3. Here we report a generally applicable platform for autonomous protein engineering that integrates machine learning and large language models with biofoundry automation to eliminate the need for human intervention, judgement, and domain expertise. Requiring only an input protein sequence and a quantifiable way to measure fitness, this autonomous platform can be applied to engineer virtually any protein. As a proof of concept, we engineered Arabidopsis thaliana halide methyltransferase (AtHMT)4 for a 90-fold improvement in substrate preference and 16-fold improvement in ethyltransferase activity, along with developing a Yersinia mollaretii phytase (YmPhytase)5,6 variant with 26-fold improvement in activity at neutral pH. This was accomplished in four rounds over four weeks, while requiring construction and characterization of fewer than a total of 500 variants for each enzyme. This platform for autonomous experimentation paves the way for rapid advancements across diverse industries, from medicine and biotechnology to renewable energy and sustainable chemistry.

synthetic biology↗

bioRxiv2024

Enhancing lipid production in plant cells through high-throughput genome editing and phenotyping via a scalable automated pipeline

Plant bioengineering is a time-consuming and labor-intensive process, with no guarantee of achieving the desired trait. Here we report a fast, automated, scalable, high-throughput pipeline for plant bioengineering (FAST-PB). FAST-PB achieves gene cloning, genome editing, and product characterization by integrating automated biofoundry engineering of callus and protoplast cells with single cell matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). We first demonstrate that FAST-PB can streamline the Golden Gate cloning process, with the capacity to construct 96 vectors in parallel. To prove the concept, using FAST-PB, we first found that PEG2050 significantly increases transfection efficiency by over 45%. To validate the pipeline, we established a reporter-gene-free method for CRISPR editing via mutation of HCF136, affecting cellular chlorophyll fluorescence. Next, we applied this pipeline for lipid production and found that diverse lipids were significantly enhanced up to sixfold through introducing multi-gene cassettes via CRISPR activation, and regenerated plant using this platform. Lastly, we harnessed FAST-PB to achieve high-throughput single-cell lipid profiling through the integration of MALDI-MS with the biofoundry, and differentiated engineered and unengineered cells using the single-cell lipidomics. These innovations massively increase the throughput of synthetic biology, genome editing, and metabolic engineering, and change what is possibly using single-cell metabolomics in plants.

synthetic biology↗

bioRxiv2024

Inhibition of Cyp1a Protects Mice against Anthracycline Cardiomyopathy

BackgroundAnthracyclines such as doxorubicin (Dox) are highly effective anti-tumor agents, but their use is limited by dose-dependent cardiomyopathy and heart failure. Our laboratory previously reported that induction of cytochrome P450 family 1 (Cyp1) enzymes contributes to acute Dox cardiotoxicity in zebrafish and in mice, and that potent Cyp1 inhibitors prevent cardiotoxicity. However, the role of Cyp1 enzymes in chronic Dox cardiomyopathy, as well as the mechanisms underlying cardioprotection associated with Cyp1 inhibition, have not been fully elucidated. MethodsThe Cyp1 pathway was evaluated using a small molecule Cyp1 inhibitor in wild-type (WT) mice, or Cyp1-null mice (Cyp1a1/1a2-/-, Cyp1b1-/-, and Cyp1a1/1a2/1b1-/-). Low-dose Dox was administered by serial intraperitoneal or intravenous injections, respectively. Expression of Cyp1 isoforms was measured by RT-qPCR, and myocardial tissue was isolated from the left ventricle for RNA sequencing. Cardiac function was evaluated by transthoracic echocardiography. ResultsIn WT mice, Dox treatment was associated with a decrease in Cyp1a2 and increase in Cyp1b1 expression in the heart and in the liver. Co-treatment of WT mice with Dox and the novel Cyp1 inhibitor YW-130 protected against cardiac dysfunction compared to Dox treatment alone. Cyp1a1/1a2-/- and Cyp1a1/1a2/1b1-/- mice were protected from Dox cardiomyopathy compared to WT mice. Male, but not female, Cyp1b1-/- mice had increased cardiac dysfunction following Dox treatment compared to WT mice. RNA sequencing of myocardial tissue showed upregulation of Fundc1 and downregulation of Ccl21c in Cyp1a1/1a2-/- mice treated with Dox, implicating changes in mitophagy and chemokine-mediated inflammation as possible mechanisms of Cyp1a-mediated cardioprotection. ConclusionsTaken together, this study highlights the potential therapeutic value of Cyp1a inhibition in mitigating anthracycline cardiomyopathy.

molecular biology↗

bioRxiv2024

Metabolic Vulnerabilities of Temozolomide-Resistant Glioblastoma Cells: Implications for Targeted Therapies and Overcoming Chemoresistance

Chemoresistance is a major clinical challenge in the management of glioblastoma (GB), making it difficult to achieve long-term success with traditional treatments. Therefore, there is a need for the development of novel drugs. We explored the metabolic vulnerabilities of temozolomide (TMZ)-resistant GB and their potential implications for targeted therapies. In monolayer and tumoroid cultures, we found elevated reliance on oxidative phosphorylation in TMZ-resistant cells. Notably, iron reduction in TMZ-resistant cells reduced viability and proliferation, upregulated hypoxia-inducible factor 1- (Hif1-) expression, induced autophagy, inhibited autophagic flux, and increased reactive oxygen species (ROS) generation, indicating the significance of iron in metabolic vulnerabilities of these cells. Hypoxic cells showed acquired resistance to iron chelation compared to their normoxic state, suggesting an adaptive mechanism associated to hypoxia. Viability, size, and invasion were reduced in TMZ-resistant tumoroids. Additionally, we reported IC50 for the combination of TMZ with a range of DFO and DFP, making the combination therapy a promising drug candidate to improve therapeutic treatments. TeaserCombining iron reduction and chemotherapy in drug-resistant glioblastoma cells enhances therapeutic outcomes.

cancer biology↗

bioRxiv2023

Cross-domain interactions induce community stability to benthic biofilms in proglacial streams

Cross-domain interactions are an integral part of the success of complex biofilms in natural environments. Here, we report on cross-domain interactions in biofilms of streams draining proglacial floodplains in the Swiss Alps. These streams, as a consequence of the retreat of glaciers, are characterized by multiple environmental gradients and stability that depend on the time since deglaciation. We estimate co-occurrence of prokaryotic and eukaryotic communities along this gradient and show that key community members have disproportionate effects on the stability of co-occurrence networks. The topology of the networks was similar independent of environmental gradients and stability. However, network stability was higher in the streams draining proglacial terrain that was more recently deglaciated. We find that both pro- and eukaryotes are central to the stability of these networks, which fragment upon the removal of both pro- and eukaryotic taxa. These keyplayers are not always abundant, suggesting an underlying functional component to their contributions. Thus, we show that there is a key role played by individual taxa in determining microbial community stability of glacier-fed streams.

microbiology↗

bioRxiv2021

Expression of CD49f defines subsets of human regulatory T cells with divergent transcriptional landscape and function that correlate with ulcerative colitis disease activity

ObjectiveAdoptive regulatory T cell (Treg) therapy is being trialled for treatment of different autoimmune disorders, including inflammatory bowel diseases (IBD). In-depth understanding of the biological variability of Treg in the human blood may be required to improve IBD immune monitoring and treatment strategies. MethodsThrough a combination of quantitative proteomic, multiparametric flow cytometry, RNA-sequencing data analysis and functional assays on Treg enriched from the blood of ulcerative colitis (UC) patients and healthy controls, we investigated the association between CD49f expression, Treg phenotype and function, and UC disease activity. ResultsHigh-dimensional analysis and filtering defined two distinct subsets of human Treg based on the presence or absence of CD49f with divergent transcriptional landscape and functional activities. CD49f negative Treg are enriched for functional Treg markers and present significantly increased suppressive capacity. In contrast, CD49f high Treg display a pro-inflammatory Th17-like phenotype and accumulate in the blood of UC patients. Dysregulation on CD49f Treg subsets in UC patients correlate with disease activity. ConclusionOverall our findings uncover the importance of CD49f expression on Treg in physiological immunity and in pathological autoimmunity.

immunology↗

bioRxiv2020

High-Resolution Metabolic Mapping of the Cerebellum Using a Reduced Field of View (Zoomed) Magnetic Resonance Spectroscopic Imaging

PurposeThe human cerebellum plays an important role in functional activity cerebrum which is ranging from motor to cognitive activities since due to its relaying role between spinal cord and cerebrum. The cerebellum poses many challenges to magnetic resonance spectroscopic imaging (MRSI) due to the caudal location, the susceptibility to physiological artifacts and partial volume artifact due to its complex anatomical structure. Thus, in present study, we propose a high-resolution MRSI acquisition scheme for the cerebellum. MethodsA zoomed or reduced-field of view (rFOV) metabolite-cycled full-intensity magnetic resonance spectroscopic imaging (MRSI) at 3T with a nominal resolution of 62.5 L was developed. Single-slice rFOV MRSI data were acquired from the cerebellum of 5 healthy subjects with a nominal resolution of 2.5{square}x{square}2.5{square}mm2 in 9{square}minutes 36. Spectra were quantified with LCModel. A spatially unbiased atlas template of the cerebellum was used for analyzing metabolite distributions in the cerebellum. ResultsThe high quality of the achieved spectra enabled to generate a high-resolution metabolic map of total N-acetylaspartate, total creatine, total choline, glutamate+glutamine and myo-inositol with Cramer-Rao lower bounds below 50%. A spatially unbiased atlas template of the cerebellum-based region of interest (ROIs) analysis resulted in spatially dependent metabolite distributions in 9 ROIs. The group-averaging across subjects in the Montreal Neurological Institute-152 template space allowed to generate a very high-resolution metabolite maps in the cerebellum. ConclusionThese findings indicate that very high-resolution metabolite probing of cerebellum is feasible using rFOV or zoomed MRSI at 3T.

neuroscience↗

bioRxiv2018

A Mathematical Modeling Approach to The Cort-Fitness Hypothesis

The Cort-Fitness hypothesis has generated much interest from investigators integrating field endocrinology with evolutionary biology, ecology, and conservation. The hypothesis was developed on the assumption that if glucocorticoid levels increase with environmental challenges and fitness decreases with environmental challenges, then there should be a negative relationship between glucocorticoid levels and fitness. However, studies across diverse taxa have found that the relationship between glucocorticoid levels and fitness is not consistent: some studies show a positive relationship, others negative, and some show no correlation. Hence, support for the hypothesis is not consistent and thus a deeper understanding of the mechanisms underlying the relationship between glucocorticoid levels, environmental pressures, and fitness is needed. We propose a mathematical model representing the links between glucocorticoid levels, environmental challenges, and fitness. Our model explores how variation in the predictability and intensity of environmental challenges, reproductive strategies, and fitness metrics can all contribute to the variability observed in empirical tests of the Cort-Fitness hypothesis. We provide qualitative results showing the Cort-Fitness relationship for different environmental scenarios and discuss how the model can be used to inform future Cort-Fitness studies.

animal behavior and cognition↗

bioRxiv2018

Risk Factors for Fouling Biomass: Evidence from Small Vessels in Australia

Invasive marine non-indigenous species are a major threat to marine biodiversity and marine related industries. Recreational vessels are recognised as an important vector of non-indigenous species translocation, particularly the secondary translocation of species domestically.\n\nThis paper reports on a novel application of multilevel modelling and multiple imputation to biomass samples gathered from the hull and other external surfaces of recreational yachts and fishing vessels in order to quantify the relationship between the wet biomass of biofouling and vessel-level characteristics. Unsurprisingly, we find that the number of days since the vessel was last cleaned was strongly related to the wet weight of biomass. The number of days since the vessel was last used was also related to the wet weight of biomass, yet differed depending on the vessel type. Similarly, the median number of trips undertaken by the vessel was related to the wet weight of biomass, and varied according to the type of antifouling paint used by the vessel. The relationship between vessel size, as measured by hull surface area, and wet weight biomass per sample unit area was not significant.\n\nIn order to reduce the international and domestic spread of invasive species, small vessel owners should use an appropriate type of antifouling paint that suits their vessels operational profile, and be encouraged to maintain a maintenance

ecology↗