Biology subjects

Brown, E.

Publications and source records attributed to Brown, E..

At least 19 recordsLinked to original sources

bioRxiv2026

Response to SMAC mimetics in patient-derived models of triple-negative breast cancer relies on RIPK1 activation

Triple-negative breast cancer (TNBC) is a highly metastatic and aggressive breast cancer subtype with limited targeted treatment options. Second mitochondrial activator of caspases (SMAC) mimetics represent a promising therapeutic class for TNBC patients, although they have yet to be approved clinically. Screening a repository of 69 breast cancer patient-derived xenograft organoid (PDxO) lines revealed that approximately 39% were sensitive to the SMAC mimetic birinapant. To define the basis of resistance in lines that did not respond, we compared birinapant-sensitive (S-) and de novo resistant (R-) TNBC PDxOs. Birinapant caused degradation of inhibitor of apoptosis proteins (IAPs) in both groups but failed to induce cell death in R-PDxOs. We determined that R-PDxOs were susceptible to certain types of apoptosis but were specifically resistant to TNFR-mediated cell death, an integral pathway involved in SMAC mimetic response. Moreover, RIPK1 activation via autophosphorylation, a critical event in TNFR-mediated apoptosis, was absent in both R-PDxOs and PDxOs selected for acquired birinapant resistance. Pharmacologic inhibition or degradation of RIPK1 protected S-PDxOs from birinapant-induced cell death, demonstrating that RIPK1 activation is required for response in TNBC. To investigate how to restore RIPK1 activation in R-PDxOs, we discovered that HOIP, an E3 ubiquitin ligase that inhibits RIPK1 autophosphorylation, was more abundant in R-PDxOs compared to S-PDxOs. Inhibiting HOIP activity restored RIPK1 autophosphorylation and sensitized a subset of R-PDxOs to birinapant. Together, these findings identify impaired RIPK1 autophosphorylation as a mechanism of birinapant resistance in TNBC and suggest that RIPK1 activation state may distinguish SMAC mimetic-sensitive and -resistant tumors. Targeting HOIP may provide a strategy to overcome resistance in selected tumors.

cancer biology↗

bioRxiv2026

A phosphorylation-dependent mechanism controls splice variant-specific S-palmitoylation of cardiac Kv4.3

S-palmitoylation modulates the activity of many cardiac ion channels, yet the upstream signals that control this post-translational modification (PTM) are poorly defined. Here we identify a phosphorylation-dependent mechanism that governs splice variant-specific S-palmitoylation of the potassium channel Kv4.3. Using Acyl-RAC in native tissue and heterologous cells, we map palmitoylation to C546/547 in the intrinsically disordered Kv4.3 C-terminal tail. The short Kv4.3 splice variant Kv4.3S is [~]2.5-3-fold more palmitoylated than the long splice variant Kv4.3L, and systematic mutagenesis localises the dominant inhibitory determinant of Kv4.3 S-palmitoylation to residues 488-498 within the Kv4.3L-specific splice insert. KChIP2.1 promotes accumulation of a post-translationally modified Kv4.3 species that is selectively S-palmitoylated, and nanobody-targeted dephosphorylation removes this species and reduces Kv4.3 S-palmitoylation. Phos-tag electrophoresis and C-terminal truncation mapping identify S538 as the principal phosphorylation site enabling Kv4.3 S-palmitoylation; mutation of S538 markedly reduces formation of the palmitoylation-competent species. TurboID proximity-labelling and biochemical assays indicate that phosphorylation enhances recruitment of Kv4.3 to zDHHC5. Acute kinase inhibition rapidly eliminates phosphorylation but only gradually reduces palmitoylation, revealing temporal uncoupling between these PTMs. Functionally, non-palmitoylatable Kv4.3S exhibits larger peak currents, faster inactivation, and a left-shifted activation curve, consistent with palmitoylation limiting channel function and modulating gating transitions. Together, these findings identify phosphorylation of S538 as a priming modification that licenses Kv4.3 S-palmitoylation at C546/547, explain splice-variant differences in S-palmitoylation, and define a PTM cascade that tunes Kv4.3 channel behaviour.

cell biology↗

bioRxiv2026

BAYESIAN STATE-SPACE MODEL FOR JOINT INFERENCE OF OSCILLATORY DYNAMICS AND POINT-PROCESS COUPLING

Under a range of behavioral and physiological conditions, spike times and local field potential (LFP) oscillations exhibit phase coupling within specific frequency bands. Classical measures such as spike-field coherence (SFC) and the phase-locking value (PLV) quantify this coupling but estimate the LFP spectrum independently of spike timing. We introduce Joint SSMT, a Bayesian state-space framework that jointly infers LFP spectrograms and spike-field coupling strength. The model treats narrowband LFP activity as a latent process evolving in continuous time, with spike trains linked to the complex spectral state through a Bernoulli-logistic model. In simulations, Joint SSMT accurately recovers coupling strength, denoises the spectrogram, and uses spike timing to resolve fine temporal structure in the LFP. Applied to propofol anesthesia data, the model identifies coupling at a specific slow-oscillation frequency where SFC and PLV report only broad low-frequency coupling. We extend Joint SSMT to trial-structured experiments and apply it to primate recordings during an associative learning task, revealing frequency-specific coupling in hippocampus and prefrontal cortex. We also derive closed-form expressions for SFC and PLV as functions of the generative model parameters. Across simulations and two primate datasets, Joint SSMT provides more frequency-specific coupling estimates with principled uncertainty quantification than classical PLV and SFC.

neuroscience↗

bioRxiv2026

Co-limitation by stable, dynamic and directional habitat features shapes climate vulnerability in an alpine specialist

Alpine ecosystems are among the most climate sensitive on Earth, yet logistical challenges and detection biases often impede robust assessment of alpine dependent species. We investigated habitat associations and density patterns of the Sierra Nevada subspecies of the Gray-crowned Rosy-Finch (Leucosticte tephrocotis dawsoni), an alpine obligate and regional endemic, over five breeding seasons from 2018 to 2022 using hierarchical distance sampling and mark-recapture distance sampling to explicitly account for imperfect detection and spatial heterogeneity. Density estimates tracked annual snowpack variation, ranging from 4.77 individuals/km{superscript 2} in a low snow year to 12.08 individuals/km{superscript 2} in a high snow year. Abundance was highest near persistent snow patches that provide foraging habitat and near cliffs that provide nesting substrate, and declined sharply above approximately 10% woody cover, with densities approaching zero beyond approximately 25%, indicating a steep ecological threshold. In contrast, the proportion of surveyed blocks with detections remained relatively stable across years. Together, these patterns indicate a three timescale co-limitation framework in which breeding habitat is shaped by static features (cliffs), dynamic annual drivers (snowpack), and longer-term directional change (woody encroachment). By linking population density to climate sensitive habitat features, this study provides a high-resolution abundance-based baseline for long term monitoring and offers a framework for evaluating climate vulnerability in alpine and other resource-limited systems. Open Research StatementData necessary to replicate the analyses and results presented in this manuscript will be archived in the Dryad Digital Repository upon acceptance, with no embargo on the material. The R code associated with this manuscript is not novel. All analyses use publicly available packages and functions without modification, including mrds (v2.3.0), unmarked (v1.2.5), tidyverse (v2.0.0), MuMIn (v1.47.5), and standard ggplot2 visualization tools. All code is properly cited within the manuscript and publicly available through CRAN. Complete analysis scripts will be archived in Dryad alongside the data upon acceptance to facilitate full reproducibility of results.

ecology↗

bioRxiv2026

Mechanisms of macular oedema development and therapeutic response: An in-silico modelling study

PurposeTo investigate the interplay between biomechanics, fluid dynamics, and solute transport in Diabetic Macular Oedema (DMO) using a mechanics-based computational model, aiming to elucidate mechanisms behind variable treatment outcomes. MethodsWe developed a multiphysics model of the retina within a porous media framework. The model integrates OCT-derived geometry, vascular leakage, retinal biomechanics (including Muller cell fibre architecture), retinal pigment epithelium (RPE) function, and anti-VEGF transport. We simulated oedema development and therapeutic response by varying these parameters systematically. ResultsModel results showed that active RPE pumping is essential for maintaining retinal dehydration. Our simulations revealed a critical trade-off related to Muller cell architecture: the physiological z-shaped orientation protects against oedema but impedes anti-VEGF drug delivery to leaky vessels. In contrast, a pathological, vertical Muller cell alignment increases oedema susceptibility but allows for a faster therapeutic response due to improved drug diffusion. ConclusionsMuller cell orientation presents a trade-off between biomechanical protection and therapeutic efficacy, offering a novel mechanistic explanation for the variable patient responses to anti-VEGF therapy observed clinically. This in-silico framework is a powerful tool for dissecting DMO pathophysiology and has the potential to guide the development of personalised treatment strategies.

biophysics↗

bioRxiv2026

Direct effects on endothelial cells are essential for perivascular cell (pericyte)-dependent amplification of orthoflavivirus NS1-mediated microvascular leakage

The emerging tick-borne flaviviruses Alkhumra haemorrhagic fever virus (AHFV) and Kyasanur Forest disease virus (KFDV) can cause severe haemorrhagic disease, yet the mechanisms driving this vascular leakage remain unclear. The microvascular capillaries and post-capillary venules affected during orthoflavivirus-related haemorrhagic disease consist of an endothelial cell barrier ensheathed in and supported by perivascular cells (pericytes), and we recently identified a critical role for pericytes in amplifying dengue microvascular dysfunction. The orthoflavivirus non-structural protein 1 (NS1) has been implicated in endothelial dysfunction and vascular leakage for several tick- and mosquito-borne flaviviruses. Here, we examined whether NS1 from AHFV and KFDV disrupts microvascular barrier integrity in pericyte-endothelial cell cocultures. We found AHFV and KFDV NS1 to disrupt the ability of pericytes to support endothelial cell function, which is required for maintenance of the microvascular barrier. However, under our experimental conditions, we detected no endothelial hyperpermeability upon NS1 treatment in endothelial cells cultured alone or in combination with pericytes. Our findings suggest that the concomitant impairment of both endothelial cell and pericyte function may be required for NS1 to induce microvascular hyperpermeability. Our work highlights the potentially synergistic effects of microvascular cells during orthoflavivirus haemorrhage and emphasises the need for further work into the mechanisms of vascular dysfunction during AHFV and KFDV infection.

microbiology↗

bioRxiv2025

Functional interrogation uncovers a critical role for a high-plasticity cell state in lung adenocarcinoma

Plasticity--the ability of cells to undergo phenotypic transitions--drives cancer progression and therapy resistance1-3. To date, strategies targeting cancer plasticity have not advanced to the clinic due to a lack of fundamental understanding of the underlying mechanisms. Recent studies have suggested that plasticity in solid tumors is concentrated in a minority subset of cancer cells4-6, yet functional studies interrogating this high plasticity cell state (HPCS) in situ are lacking. Here, we developed mouse models enabling detection, longitudinal lineage tracing, and ablation of the HPCS in autochthonous lung tumors in vivo. Using lineage tracing, we uncover the HPCS cells are dedifferentiated but possess high capacity for cell state transitions, giving rise to both early neoplastic (differentiated) and advanced lung cancer cell states in situ. Longitudinal lineage tracing using secreted luciferases reveals HPCS-derived cells harbor high capacity for growth when compared to bulk cancer cells or another defined cancer cell state with features of differentiated lung epithelium. Suicide gene-mediated ablation of the HPCS in early neoplasias abrogates tumor progression. Ablating HPCS cells in established tumors by suicide gene or HPCS-directed CAR T cells robustly reduces tumor burden, whereas ablation of a differentiated lung cancer cell state had no effect. We further demonstrate that the HPCS gives rise to therapy-resistant cell states, whereas ablation of the HPCS abrogates resistance to chemotherapy and oncoprotein-targeted therapy. Interestingly, an HPCS-like state is ubiquitous in regenerating epithelia and in carcinomas of multiple other tissues, revealing a convergence of plasticity programs. Our work establishes the HPCS as a critical hub enabling reciprocal transitions between cancer cell states, including acquisition of states adapted to cancer therapies. Targeting the HPCS in lung cancer and in other carcinomas may suppress cancer progression and eradicate treatment resistance.

cancer biology↗

bioRxiv2024

Reducing oxidative stress improves ex vivo polymer-based human haematopoietic stem and progenitor cell culture and gene editing

Self-renewing multipotent haematopoietic stem cells (HSCs) have the unique capacity to stably regenerate the entire blood and immune systems following transplantation. HSCs are used clinically to reconstitute a healthy blood system in patients suffering from a range of haematological diseases. However, HSCs are very rare and have been challenging to grow ex vivo, which has hampered efforts to collect large numbers of HSCs for both basic research and clinical therapies. Polymer-based culture conditions have recently been developed to support expansion of mouse and human haematopoietic stem and progenitor cells (HSPCs). While mouse HSPCs expanded rapidly in polymer-based cultures, growth speeds for human HSPCs in polymer-based cultures was limited to [~]70-fold over 4-weeks. Here we have found that reducing oxidative stress improves human HSPC growth in these conditions. We describe an optimised culture condition that improves growth to 250-1400-fold over 4-weeks through reducing oxygen tension and the addition of antioxidants. These conditions also enable efficient gene editing in these polymer-based cultures. We envision these improved culture conditions will support a range of research into human HSPC biology and provide a platform for clinical-scale HSPC expansion and gene editing.

cell biology↗

bioRxiv2024

Human placental organoids as a model to probe early gestation maternal immune dynamics

During pregnancy, the human placenta establishes tolerance toward fetal allogeneic tissue, where specialized trophoblast subtypes play a complex role in local and peripheral immunomodulation. However, due to inadequate models to study the early gestation of the human placenta, each trophoblast subtypes role in modulating the maternal immune response has remained elusive. Here, we derived human placental organoids from early gestation trophoblast stem cells to (1) identify patterns of immunomodulatory protein expression by trophoblast subtype and (2) evaluate the effects of the placental organoid secretome on immune cell activation and regulation. We show that the three primary trophoblast phenotypes had distinct influences on immune cell phenotype and activation and that three-dimensional culture significantly alters trophoblast immunomodulation relative to traditional two-dimensional trophoblast culture.

bioengineering↗

bioRxiv2024

Student experiences with an interactive 3D immersive biotechnology simulation and its impact on motivational beliefs

The development and use of virtual laboratories to augment traditional in-person skills training continues to grow. Virtual labs have been implemented in a number of diverse educational settings, which have many purported benefits including their adaptability, accessibility, and repeatability. However, few studies have evaluated the impact of virtual laboratories outside of academic achievement and skills competencies, especially in biotechnology. In this study, an interdisciplinary team of content experts, video game researchers, instructional designers, and assessment experts developed a 3D immersive simulation designed to teach novice scientists the technical skills necessary to perform sterile mammalian cell culture technique. Unique to the simulation development process is the recreation of an immersive experience through the capture of details in the real-world lab where participants have the freedom of choice in their actions, while receiving immediate feedback on their technical skills as well as procedural execution. However, unlike an in-person laboratory course, students are able to iterate and practice their skills outside of class time and learn from their mistakes. Using a mixed-methods study design, over the course of two semesters we evaluated student attitudes of the simulation and their science motivational beliefs including self-efficacy and science identity after engaging with the simulation prior to the physical laboratory. Our results show that students science identity remained unchanged while their science self-efficacy increased. Furthermore, students had positive perceptions of the benefits of the virtual simulation. These data suggest that the virtual cell culture simulation can be a useful pedagogical training tool to bolster students motivational beliefs that is both accessible and easy to implement.

scientific communication and education↗

bioRxiv2024

USP50 suppresses alternative RecQ helicase use and deleterious DNA2 activity during replication.

Mammalian DNA replication employs several RecQ DNA helicases to orchestrate the faithful duplication of genetic information. Helicase function is often coupled to the activity of specific nucleases, but how helicase and nuclease activities are co-directed is unclear. Here we identify the inactive ubiquitin-specific protease, USP50, as a ubiquitin-binding and chromatin-associated protein required for ongoing replication, fork restart, telomere maintenance and cellular survival during replicative stress. USP50 supports WRN:FEN1 at stalled replication forks, suppresses MUS81-dependent fork collapse and restricts double-strand DNA breaks at GC-rich sequences. Surprisingly we find that cells depleted for USP50 and recovering from a replication block exhibit increased DNA2 and RECQL4 foci and that the defects in ongoing replication, poor fork restart and increased fork collapse seen in these cells are mediated by DNA2, RECQL4 and RECQL5. These data define a novel ubiquitin-dependent pathway that promotes the balance of helicase: nuclease use at ongoing and stalled replication forks.

cancer biology↗

bioRxiv2023

Gamma sensory stimulation and effects on the brain

Findings by the Tsai lab and others 1-8 demonstrate that 40 Hz frequency sensory stimulation induces electrophysiological responses and attenuates pathology in mouse models of Alzheimers disease (AD). A recent study in Nature Neurosciene 9 concluded that the stimulation does not affect endogenous gamma oscillations or amyloid burden. We welcome research investigating 40 Hz sensory stimulation, and the article by Soula et al enhances our understanding of the brains electrophysiological response to 40Hz. However, we respectfully suggest that the data in Soula et al are consistent with a neuronal response to 40 Hz, which we further support with new data in humans. Moreover we contend the non-significant effects on amyloid are due to technical limitations of the study.

neuroscience↗

bioRxiv2023

Physics-informed deep generative learning for quantitative assessment of the retina

Disruption of retinal vasculature is linked to various diseases, including diabetic retinopathy and macular degeneration, leading to vision loss. We present here a novel algorithmic approach that generates highly realistic digital models of human retinal blood vessels based on established biophysical principles, including fully-connected arterial and venous trees with a single inlet and outlet. This approach, using physics-informed generative adversarial networks (PI-GAN), enables the segmentation and reconstruction of blood vessel networks that requires no human input and out-performs human labelling. Our findings highlight the potential of PI-GAN for accurate retinal vasculature characterization, with implications for improving early disease detection, monitoring disease progression, and improving patient care.

biophysics↗

bioRxiv2023

Microengineering 3D Collagen Matrices with Tumor-Mimetic Gradients in Fiber Alignment

In the tumor microenvironment (TME), collagen fibers facilitate tumor cell migration through the extracellular matrix. Previous studies have focused on studying the responses of cells on uniformly aligned or randomly aligned collagen fibers. However, the in vivo environment also features spatial gradients in alignment, which arise from the local reorganization of the matrix architecture due to cell-induced traction forces. Although there has been extensive research on how cells respond to graded biophysical cues, such as stiffness, porosity, and ligand density, the cellular responses to physiological fiber alignment gradients have been largely unexplored. This is due, in part, to a lack of robust experimental techniques to create controlled alignment gradients in natural materials. In this study, we image tumor biopsy samples and characterize the alignment gradients present in the TME. To replicate physiological gradients, we introduce a first-of-its-kind biofabrication technique that utilizes a microfluidic channel with constricting and expanding geometry to engineer 3D collagen hydrogels with tunable fiber alignment gradients that range from sub-millimeter to millimeter length scales. Our modular approach allows easy access to the microengineered gradient gels, and we demonstrate that HUVECs migrate in response to the fiber architecture. We provide preliminary evidence suggesting that MDA-MB-231 cell aggregates, patterned onto a specific location on the alignment gradient, exhibit preferential migration towards increasing alignment. This finding suggests that alignment gradients could serve as an additional taxis cue in the ECM. Importantly, our study represents the first successful engineering of continuous gradients of fiber alignment in soft, natural materials. We anticipate that our user-friendly platform, which needs no specialized equipment, will offer new experimental capabilities to study the impact of fiber-based contact guidance on directed cell migration.

bioengineering↗

bioRxiv2022

TORC1 is an essential regulator of nutrient-dependent differentiation in Leishmania

Leishmania parasites undergo differentiation between various proliferating and non-dividing forms to adapt to changing host environments. The mechanisms that link environmental cues with the parasites developmental changes remain elusive. Here, we report that Leishmania TORC1 is a key environmental sensor for parasite differentiation in the sand fly-stage promastigotes and for replication of mammalian-stage amastigotes. We show that Leishmania RPTOR1, interacts with TOR1 and LST8. We investigate TORC1 function by conditional deletion of RPTOR1, where under nutrient rich conditions RPTOR1 depletion results in decreased protein synthesis and growth, G1 cell cycle arrest and premature differentiation from proliferative promastigotes to non-dividing mammalian-infective metacyclic forms. These parasites cannot develop into proliferative amastigotes in the mammalian host, or respond to nutrients to differentiate to proliferative retroleptomonads, which are required for their blood-meal induced amplification in sand flies and enhanced mammalian infectivity. RPTOR1-dependent TORC1 functionality represents a critical mechanism for driving parasite growth and proliferation.

cell biology↗

bioRxiv2022

Inferring transmission routes for foot-and-mouth disease virus within a cattle herd using approximate Bayesian computation

To control an outbreak of an infectious disease it is essential to understand the different routes of transmission and how they contribute to the overall spread of the pathogen. With this information, policy makers can choose the most efficient methods of detection and control during an outbreak. Here we demonstrate a method for assessing the contribution of different routes of transmission using approximate Bayesian computation with sequential Monte Carlo sampling (ABC-SMC). We apply this to infer parameters of an individual based model of within-herd transmission of foot-and-mouth disease virus (FMDV), incorporating transmission through direct contact and via environmental contamination. Additionally, we use ABC-SMC for model selection to assess the plausibility of either transmission route alone being responsible for all infections. We show that direct transmission likely contributes the majority of infections during an outbreak of FMD but there is an accumulation of environmental contamination that can cause infections within a farm and also have the potential to spread between farms via fomites.

zoology↗

bioRxiv2022

Novelty detection and multiple timescale integration drive Drosophila orientation dynamics in temporally diverse olfactory environments

To survive, insects must effectively navigate odors plumes to their source. In natural plumes, turbulent winds break up smooth odor regions into disconnected patches, so navigators encounter brief bursts of odor interrupted by bouts of clean air. The timing of these encounters plays a critical role in navigation, determining the direction, rate, and magnitude of insects orientation and speed dynamics. Still, disambiguating the specific role of odor timing from other cues, such as spatial structure, is challenging due to natural correlations between plumes temporal and spatial features. Here, we use optogenetics to isolate temporal features of odor signals, examining how the frequency and duration of odor encounters shape the navigational decisions of freely-walking Drosophila. We find that fly angular velocity depends on signal frequency and intermittency - fraction of time signal can be detected - but not directly on durations. Rather than switching strategies when signal statistics change, flies smoothly transition between signal regimes, by combining an odor offset response with a frequency-dependent novelty-like response. In the latter, flies are more likely to turn in response to each odor hit only when the hits are sparse. Finally, the upwind bias of individual turns relies on a filtering scheme with two distinct timescales, allowing rapid and sustained responses in a variety of signal statistics. A quantitative model incorporating these ingredients recapitulates fly orientation dynamics across a wide range of environments.

animal behavior and cognition↗

bioRxiv2022

Cysteine post-translational modifications regulate protein interactions of caveolin-3

Caveolae are small flask-shaped invaginations of the surface membrane which are proposed to recruit and co-localise signalling molecules. The distinctive caveolar shape is achieved by the oligomeric structural protein caveolin, of which three isoforms exist. Aside from the finding that caveolin-3 is specifically expressed in muscle, functional differences between the caveolin isoforms have not been rigorously investigated. Caveolin-3 is relatively cysteine-rich compared to caveolins 1 and 2, so we investigated its cysteine post-translational modifications. We find that caveolin-3 is palmitoylated at 6 cysteines and becomes glutathiolated following redox stress. We map the caveolin-3 palmitoylation sites to a cluster of cysteines in its C terminal membrane domain, and the glutathiolation site to an N terminal cysteine close to the region of caveolin-3 proposed to engage in protein interactions. Glutathiolation abolishes caveolin-3 interaction with heterotrimeric G protein alpha subunits. Our results indicate that a caveolin-3 oligomer contains up to 66 palmitates, compared to up to 33 for caveolin-1. The additional palmitoylation sites in caveolin-3 therefore provide a mechanistic basis by which caveolae in smooth and striated muscle can possess unique phospholipid and protein cargoes. These unique adaptations of the muscle-specific caveolin isoform have important implications for caveolar assembly and signalling.

cell biology↗