Biology subjects

Cox, S.

Publications and source records attributed to Cox, S..

At least 19 recordsLinked to original sources

bioRxiv2025

The 9-1-1 complex protects ssDNA gaps in BRCA2-deficient cancer

Single-stranded DNA (ssDNA) gaps are a hallmark of BRCA-deficient cells, yet the mechanisms that safeguard these lesions remain unclear. Through a genome-wide CRISPR screen, we identified the RAD9A-HUS1-RAD1 (9-1-1) complex as essential for the survival of BRCA2-deficient cells through an ATR-independent mechanism. Loss of 9-1-1 in this context leads to the accumulation of PRIMPOL-dependent gaps that fail to undergo post-replicat ive repair, resulting in pathological expansion and increased DNA damage. This instability is driven by excessive EXO1-mediated degradation, as EXO1 depletion rescues the phenotype. We further demonstrate that the 9-1-1 complex is required for POL{zeta}-dependent gap filling. We propose a model in which ssDNA gaps, when extended beyond a critical length, become inaccessible to TLS-mediated repair and are fully reliant on homologous recombination. These findings establish the 9-1-1 complex as key regulator of ssDNA gap stability and a promising therapeutic target in BRCA2-deficient cancers. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/680950v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@56516aorg.highwire.dtl.DTLVardef@fb1ce3org.highwire.dtl.DTLVardef@25a7b2org.highwire.dtl.DTLVardef@249731_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

Image quality metrics fail to accurately represent biological information in fluorescence microscopy

Image processing methods offer the potential to improve the quality of fluorescence microscopy data, allowing for image acquisition at lower, less phototoxic illumination doses. The training and evaluation of such methods is informed and driven by full-reference image quality metrics (IQMs); however, these metrics derive from applications to natural scene images, not fluorescence microscopy images. Here we investigate the response of IQMs to common properties of fluorescence microscopy data and whether IQMs are capable of reporting the biological information content of images. We find that IQM scores are biased by image content for both raw and processed microscopy data, and that improvements in IQM values reported after processing are not reliably correlated with performance in downstream analysis tasks. As common IQMs are unreliable proxies for guiding image processing developments in biological fluorescence microscopy, image processing performance should be benchmarked according to downstream analysis success.

bioinformatics↗

bioRxiv2025

Extracting biological structure and heterogeneity from the nano to the macro scale

Fluorescence microscopy is an essential tool in biology. It has revealed great variability at multiple scales, in macromolecular complexes, cells, and organisms. Understanding this variability will reveal the mechanisms by which genetically or biochemically identical systems adopt different biological states. Achieving this requires the ability to extract both the underlying biological structure and how it varies across the population. Currently the field lacks general techniques to deal with arbitrary structures and different types of variability. Here we present SQUASSH, a new convolutional neural network-based approach to freely fit structural models to fluorescence microscopy data that simultaneously quantifies variability to reveal correlations, dynamics, and systematic distortions. SQUASSH is highly versatile: it accommodates diverse imaging modalities at length scales from nm to mm. This approach opens up applications such as imaging nanoscale macromolecular structures, revealing patterns in shape changes from organelle to tissue scale, and characterizing systems biology of dynamical processes.

cell biology↗

bioRxiv2025

Orthorexia is associated with a paradoxical appetitive gastric response to unhealthy foods

Orthorexia involves an obsessive tendency towards "healthy" foods. It is a risk factor for eating disorders (anorexia nervosa and the proposed "orthorexia nervosa") and psychological distress, but its biological mechanism remains unknown. We hypothesised that this mechanism operates through the stomach: increased gastric power differentiates appetising from unappetising foods, vagal stimulation reduces food liking, and gastric proto-nausea can be evoked by disgust; all contributors to disordered eating. We used electrogastrography alongside high-density facial landmark tracking to gauge responses to minimally processed ("healthy"), highly processed ("unhealthy"), and culturally unaccepted ("disgusting") foods in a non-clinical sample (N=77). Trait orthorexia was associated with increased self-reported desire to eat healthy foods. Moreover, we found that higher orthorexia was associated with increasing disgust-related facial responses to unhealthy and disgusting foods. We then identified a relationship between food-healthiness ratings and gastric power: those who generally assigned lower healthiness ratings to foods showed higher gastric power for unhealthy than healthy foods, whereas those who expressed higher healthiness ratings showed higher gastric power for healthy over unhealthy foods. This suggests that the stomach tracks individual differences in eating preferences. Crucially, and contrary to our expectations, higher trait orthorexia was associated with a paradoxical increase in gastric power to unhealthy foods. One explanation is that higher gastric power for unhealthy foods reflects increased appetite for self-denied foods. Alternatively, orthorexia itself could be an adaptive response to exert stronger cognitive control over a pre-existing gastric appetite for unhealthy foods.

physiology↗

bioRxiv2025

C57 and DBA mouse strains express distinct cocaine avoidance phenotypes in an operant runway independent of differences in striatal dynorphin-enkephalin balance

RationaleHuman self-report data and rodent models indicate cocaine can induce negative affect, which may impact future cocaine use severity. Despite this, understanding of the neurobiology driving cocaine avoidance is limited. Within the striatum, the opioid peptides enkephalin and dynorphin are associated with cocaine reward and aversion, respectively. Additionally, striatal dynorphin signaling resulting from stress or cocaine withdrawal acts as a negative reinforcer to increase cocaine seeking. ObjectivesWe validated the use of the cocaine self-administration runway in mice to measure the development of cocaine avoidance and tested whether mice with higher relative striatal dynorphin-to-enkephalin expression are resistant to developing cocaine avoidance. MethodsCocaine avoidance in the self-administration runway was measured in two inbred mouse strains, C57BL/6J and DBA/2J, known to have opposite striatal dynorphin/enkephalin milieus, and in mice lacking enkephalin selectively from striatal medium spiny neurons (D2-PenkKO). ResultsBoth inbred strains developed cocaine avoidance, though they expressed it differently. Across training, DBA/2J mice increased their latency to self-administer cocaine, and C57BL/6J increased the number of retreats away from the cocaine-paired goal box. D2-PenkKOs developed similar cocaine avoidance compared to littermate controls. ConclusionsMice develop avoidance to self-administer cocaine in the runway across a range of strains. Pre-existing strain differences in the striatal dynorphin/enkephalin milieu, however, do not appear to alter the development of cocaine avoidance, and striatal enkephalin is not necessary for the development of cocaine avoidance. This suggests higher relative striatal dynorphin does not facilitate cocaine seeking by mitigating cocaine avoidance.

neuroscience↗

bioRxiv2024

A hybrid computational model of cancer spheroid growth with ribose-induced collagen stiffening

Metastasis, the leading cause of death in cancer patients, arises when cancer cells disseminate from a primary solid tumour to distant organs. Growth and invasion of the solid tumour often involve collective cell migration, which is profoundly influenced by cell-cell interactions and the extracellular matrix (ECM). The ECMs biochemical composition and mechanical properties, such as stiffness, regulate cancer cell behaviour and migration dynamics. Mathematical modelling serves as a pivotal tool for studying and predicting these complex dynamics, with hybrid discrete-continuous models offering a powerful approach by combining agent-based representations of cells with continuum descriptions of the surrounding microenvironment. In this study, we investigate the impact of ECM stiffness, modulated via ribose-induced collagen cross-linking, on cancer spheroid growth and invasion. We employed a hybrid discrete-continuous model implemented in PhysiCell to simulate spheroid dynamics, successfully replicating three-dimensional in vitro experiments. The model incorporates detailed representations of cell-cell and cell-ECM interactions, ECM remodelling, and cell proliferation. Our simulations align with experimental observations of two breast cancer cell lines, non-invasive MCF7 and invasive HCC1954, under varying ECM stiffness conditions. The results demonstrate that increased ECM stiffness due to ribose-induced cross-linking inhibits spheroid invasion in invasive cells, whereas non-invasive cells remain largely unaffected. Furthermore, our simulations show that higher ECM degradation by the cells not only enables spheroid growth and invasion but also facilitates the formation of multicellular protrusions. Conversely, increasing the maximum speed that cells can reach due to cell-ECM interactions enhances spheroid growth while promoting single-cell invasion. This hybrid modelling approach enhances our understanding of the interplay between cancer cell migration, proliferation, and ECM mechanical properties, paving the way for future studies incorporating additional ECM characteristics and microenvironmental conditions.

systems biology↗

bioRxiv2024

Force-transducing molecular ensembles at growing microtubule tips control mitotic spindle size

Mitotic spindle is a complex bipolar cellular structure that ensures chromosomes segregation between dividing cells. Correct spindle size is required for the accurate segregation and successful passing of genomes to the newly formed cells. The spindle size is believed to be controlled by mechanical forces generated by molecular motors and non-motor proteins acting in the spindle microtubule overlaps. However, how forces generated by individual proteins enable bipolar spindle organization is not well understood. Here, we developed tools to measure contributions of individual molecules to this force balance. We show that microtubule tip-trackers act synergetically at microtubule tips with minus-end directed motors to produce a system that can generate both pushing and pulling forces. We show that this system harnesses forces generated by growing tips of spindle microtubules and provides unique contribution to the force balance distinct from other force generators because it acts at microtubule tips rather than in microtubule overlaps. We show that this system alone can establish stable bipolar organization in vitro and in mitotic spindles in human cells. Our results pave the way for understanding how mechanical forces in spindles can be fine-tuned to control the fidelity of chromosome segregation.

biophysics↗

bioRxiv2023

Characterisation and correction of polarisation effects in fluorescently labelled fibres

Many biological structures take the form of fibres and filaments, and quantitative analysis of fibre organisation is important for understanding their functions in both normal physiological conditions and disease. In order to visualise these structures, fibres can be fluorescently labelled and imaged, with specialised image analysis methods available for quantifying the degree and strength of fibre alignment. Here we show that fluorescently labelled fibres can display polarised emission, with the strength of this effect varying depending on structure and fluorophore identity. This can bias automated analysis of fibre alignment and mask the true underlying structural organisation. We present a method for quantifying and correcting these polarisation effects without requiring polarisation resolved microscopy, and demonstrate its efficacy when applied to images of fluorescently labelled collagen gels, allowing for more reliable characterisation of fibre microarchitecture.

cell biology↗

bioRxiv2023

Sox9 Regulates Extracellular Matrix Stiffness during Vascular Smooth Muscle Cell ageing by increased secretion of LH3 via extracellular vesicles.

RationaleVascular calcification and increased extracellular matrix (ECM) stiffness are hallmarks of vascular ageing. Sox9 (SRY-Box Transcription Factor 9) is a master regulator of chondrogenesis, also expressed in the vasculature, that has been implicated in vascular smooth muscle cell (VSMC) osteo-chondrogenic conversion. ObjectiveHere, we investigated the relationship between vascular ageing, calcification and Sox9-driven ECM regulation in VSMCs. Methods and ResultsImmunohistochemistry in human aortic samples showed that Sox9 was not spatially associated with vascular calcification but correlated with the senescence marker p16. Analysis of Sox9 expression in vitro showed it was mechanosensitive with increased expression and nuclear translocation in senescent cells and on stiff matrices. Manipulation of Sox9 via overexpression and depletion, combined with atomic force microscopy (AFM) and proteomics, revealed that Sox9 regulates ECM stiffness and organisation by orchestrating changes in collagen expression and reducing VSMC contractility, leading to the formation of an ECM that mirrored that of senescent cells. These ECM changes promoted phenotypic modulation of VSMCs whereby senescent cells plated onto ECM synthesized from cells depleted of Sox9 returned to a proliferative state, while proliferating cells on a matrix produced by Sox9 expressing cells showed reduced proliferation and increased DNA damage, reiterating features of senescent cells. Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 3 (LH3) was identified as a Sox9 target, and key regulator of ECM stiffness. LH3 is packaged into extracellular vesicles (EVs) and Sox9 promoted EV secretion, leading to increased LH3 deposition within the ECM. ConclusionsThese findings identify cellular senescence and Sox9 as a key regulators of ECM stiffness during VSMC ageing and highlight a crucial role for ECM structure and composition in regulating VSMC phenotype. We identify a positive feedback cycle whereby cellular senescence and increased ECM stiffening promote Sox9 expression which drives further ECM modifications that act to accelerate vascular stiffening and cellular senescence.

molecular biology↗

bioRxiv2022

Neuronal, Affective, and Sensory Correlates of Targeted Helping Behavior in Male and Female Sprague Dawley Rats

Empathy is an innate ability to understand the emotional states of others along with the motivation to improve it. It has evolved over time into highly complex behaviors, the basis of which can be described using the Perception Action Model (PAM), where shared affect promotes an action that eliminates the distress of both the passive "Target" and, by extension, the active "Observer." There are myriad biological variables that may modulate empathic behavior, including sex, sensory modalities, and neural activity. In the following studies, using our labs model of social contact-independent targeted helping, we first tested whether sex differences exist in helping behavior. Next, we explored sex differences in sensory and affective signaling, including the impact of direct visualization of a distressed conspecific and the type of ultrasonic vocalizations (USV) made between animal pairs during the task. Finally, we examined the neural activity of multiple cortical and subcortical regions of interest across time during targeted helping between males and females. We show both sexes exhibit similar helping behavior, but sensory and affective signaling differs between sexes. Further, changes in neural activity exhibited distinct sex-specific patterns across time. Our results indicate sex differences are not a ubiquitous presence in targeted helping. Instead, it is likely sex differences may be a convergent phenomenon in which the behavior is similar, but the underlying biological mechanisms are distinct. These results lay the groundwork for future studies to explore the similarities and differences that drive empathic behavior in both males and females.

animal behavior and cognition↗

bioRxiv2022

Nuclear fascin regulates cancer cell survival

Fascin is an important regulator of F-actin bundling leading to enhanced filopodia assembly. Fascin is also overexpressed in most solid tumours where it supports invasion through control of F-actin structures at the periphery and nuclear envelope. Recently fascin has been identified in the nucleus of a broad range of cell types but the contributions of nuclear fascin to cancer cell behaviour remain unknown. Here we demonstrate that fascin bundles F-actin within the nucleus to support chromatin organisation and efficient DNA damage response. Fascin associates directly with phosphorylated Histone H3 leading to regulated levels of nuclear fascin to support these phenotypes. Forcing nuclear fascin accumulation through the expression of nuclear-targeted fascin-specific nanobodies or inhibition of Histone H3 kinases results in enhanced and sustained nuclear F-actin bundling leading to reduced invasion, viability and nuclear fascin-specific/driven apoptosis. These findings represent an additional important route through which fascin can support tumorigenesis and provide insight into potential pathways for targeted fascin-dependent cancer cell killing.

cancer biology↗

bioRxiv2022

Extracellular matrix assembly stress drives Drosophila central nervous system morphogenesis

The forces controlling tissue morphogenesis are attributed to cellular-driven activities and any role for extracellular matrix (ECM) is assumed to be passive. However, all polymer networks, including ECM, can theoretically develop autonomous stresses during their assembly. Here we examine the morphogenetic function of an ECM prior to reaching homeostatic equilibrium by analyzing de novo ECM assembly during Drosophila ventral nerve cord (VNC) condensation. Asymmetric VNC shortening and a rapid decrease in surface area correlate with exponential assembly of Collagen-IV (Col4) surrounding the tissue. Concomitantly, a transient developmentally-induced Col4 gradient leads to coherent long-range flow of ECM, which equilibrates the Col4 network. Finite element analysis and perturbation of Col4 network formation through the generation of dominant Col4-truncations that affect assembly, reveals that VNC morphodynamics is driven by a sudden increase in ECM-driven surface tension. These data highlight that ECM assembly stress and associated network instabilities can actively participate in tissue morphogenesis.

developmental biology↗

bioRxiv2022

Differential remodelling in small and large murine airways revealed by novel whole lung airway analysis.

Airway remodelling occurs in chronic asthma leading to increased airway smooth muscle (ASM) mass and extra-cellular matrix (ECM) deposition. Whilst extensively studied in murine airways; studies report only selected larger airways at one time point meaning the spatial distribution and resolution of remodelling are poorly understood. Here we use a new method allowing comprehensive assessment of the spatial and temporal changes in ASM, ECM and epithelium in large numbers of murine airways after allergen challenge. Using image processing to analyse 20-50 airways from a whole lung section revealed increases in ASM and ECM after allergen challenge were greater in small and large rather than intermediate airways. ASM predominantly accumulated adjacent to the basement membrane whereas ECM was distributed across the airway wall. Epithelial hyperplasia was most marked in small and intermediate airways. Post challenge, ASM changes resolved over seven days whereas ECM and epithelial changes persisted. The new method suggests large and small airways remodel differently and the long-term consequences of airway inflammation may depend more on ECM and epithelial changes than ASM. The method reduces the number of animals needed, reveals important spatial differences in remodelling and could set new analysis standards for murine asthma models.

physiology↗

bioRxiv2021

Dug: A Semantic Search Engine Leveraging Peer-Reviewed Literature to Span Biomedical Data Repositories

MotivationAs the number of public data resources continues to proliferate, identifying relevant datasets across heterogenous repositories is becoming critical to answering scientific questions. To help researchers navigate this data landscape, we developed Dug: a semantic search tool for biomedical datasets utilizing evidence-based relationships from curated knowledge graphs to find relevant datasets and explain why those results are returned. ResultsDeveloped through the National Heart, Lung, and Blood Institutes (NHLBI) BioData Catalyst ecosystem, Dug has indexed more than 15,911 study variables from public datasets. On a manually curated search dataset, Dugs total recall (total relevant results/total results) of 0.79 outperformed default Elasticsearchs total recall of 0.76. When using synonyms or related concepts as search queries, Dug (0.36) far outperformed Elasticsearch (0.14) in terms of total recall with no significant loss in the precision of its top results. Availability and ImplementationDug is freely available at https://github.com/helxplatform/dug. An example Dug deployment is also available for use at https://search.biodatacatalyst.renci.org/. Contactawaldrop@rti.org or scox@renci.org

bioinformatics↗

bioRxiv2021

Sub-diffraction error mapping for localization microscopy images

Assessing the quality of localization microscopy images is highly challenging due to difficulty in reliably detecting errors in experimental data, with artificial sharpening being a particularly common failure mode of the technique. Here we use Haar wavelet kernel analysis (HAWK), a localization microscopy data analysis method which is known to give results without artificial sharpening, to generate a reference image. This enables the mapping and quantification of this common artefact. By suppressing intensity information, we are able to map sharpening errors in a way which is not influenced by nonlinearity in the localisation imaging process. The HAWK Method for the Assessment of Nanoscopy (HAWKMAN) is a general approach which allows the reliability of localization information to be assessed.

cell biology↗

bioRxiv2020

BIOME-Preserve: A Novel Storage and Transport Medium for Preserving Anaerobic Microbiota Samples for Culture Recovery

Culture-based study design is critical to advance research into the relationship between human health and the microbiome. Traditional sample collection protocols are focused on preserving nucleic acids and metabolites and are largely inappropriate for preserving sensitive anaerobic bacteria alive for later culture recovery. Here we introduce a novel microbiome preservation kit (BIOME-Preserve) that facilitates recovery of anaerobic organisms from human stool held at room temperature. Using a combination of culture recovery and shallow whole-genome shotgun sequencing, we characterized the culturable anaerobes from fresh human stool and from human stool held in BIOME-Preserve for up to 120 hours. We recovered several species of interest to microbiome researchers, including Bifidobacterium spp., Bacteroides spp., Blautia spp., Eubacterium halii, Akkermansia muciniphila, and Faecalibacterium prausnitzii. Together, our results suggest BIOME-Preserve is practical for the collection, transport, and culture of anaerobic bacteria from human samples and can help provide the foundation for culture collections that can be used in further research and in the development of microbiome-based therapeutics. ImportanceSequencing-based protocols for studying the human microbiome have unearthed a wealth of information about the relationship between the microbiome and human health. But these microbes cannot be leveraged as therapeutic targets without culture-based studies to phenotype species of interest and to establish culture collections for use in animal models. Contrary to popular opinion, most gastrointestinal bacteria can be cultured, yet most sample collection strategies are optimized for the preservation of nucleic acids and/or metabolites only and do not take into account considerations for preserving oxygen-sensitive anaerobes and facultative anaerobes, which comprise the majority of the human gut microbiome. A human microbiome sample transport and preservation medium such as the one described here can play an important role in enabling researchers to better understand the link between the microbiome and human health and how to leverage that link through novel microbiome-based therapeutics.

microbiology↗

bioRxiv2020

Goats Given Transdermal Flunixin Meglumine Displayed Less Pain Behavior After Castration

Pain management should be utilized with castration to reduce physiological and behavioral changes. Transdermal application of drugs require less animal management and fewer labor risks, which can occur with oral administration or injections. The objective was to determine the effects of transdermal flunixin meglumine on meat goats’ behavior post-castration. Male goats (N = 18; mean body weight ± standard deviation: 26.4 ± 1.6 kg) were housed individually in pens and randomly assigned to 1 of 3 treatments: (1) castrated, dosed with transdermal flunixin meglumine; (2) castrated, dosed with transdermal placebo; and (3) sham castrated, dosed with transdermal flunixin meglumine. Body position, rumination, and head- pressing were observed for 1 h ± 10 minutes twice daily on days −1, 0, 1, 2, and 5 around castration. Each goat was observed once every 5-minutes (scan samples) and reported as percentage of observations. Accelerometers were used to measure standing, lying, and laterality (total time, bouts, and bout duration). A linear mixed model was conducted using GLIMMIX. Fixed effects of treatment, day relative to castration, and treatment*day relative to castration and random effect of date and goat nested within treatment were included. Treatment 1 goats (32.7 ± 2.8%) and treatment 2 goats (32.5 ± 2.8%) ruminated less than treatment 3 goats (47.4 ± 2.8%, P = 0.0012). Head pressing was greater on day of castration in treatment 2 goats (P < 0.001). Standing bout duration was greatest in treatment 2 goats on day 1 post-castration (P < 0.001). Lying bout duration was greatest in treatment 2 goats on day 1 post-castration compared to treatment 1 and treatment 3 goats(P < 0.001). Transdermal flunixin meglumine improved goats’ fluidity of movement post-castration and decreased head pressing, indicating a mitigation of pain behavior.View Full Text

animal behavior and cognition↗

bioRxiv2019

High-polyphenol extracts from Sorghum bicolor attenuate replication of Legionella pneumophila within RAW 264.7 macrophages

Polyphenols derived from a variety of plants have demonstrated antimicrobial activity against diverse microbial pathogens. Legionella pneumophila is an intracellular bacterial pathogen that opportunistically causes a severe inflammatory pneumonia in humans, called Legionnaires Disease, via replication within macrophages. Previous studies demonstrated that tea polyphenols attenuate L. pneumophila intracellular replication within mouse macrophages via amplification of tumor necrosis factor (TNF) production. Sorghum bicolor is a sustainable and resilient cereal crop that thrives in arid environments and is well suited to continued production in warming climates. Polyphenols derived from sorghum have anticancer and antioxidant properties, but their antimicrobial activity has not been evaluated. Here, we investigated the impact of sorghum polyphenols on L. pneumophila intracellular replication within RAW 264.7 mouse macrophages. We discovered that sorghum high-polyphenol extract (HPE) treatment attenuates L. pneumophila intracellular replication in a dose-dependent manner. Sorghum HPE did not impair bacterial replication in vitro or impact macrophage viability. Moreover, in contrast to tea polyphenols, HPE treatment impaired TNF secretion from infected macrophages. Thus, polyphenols derived from sorghum enhance macrophage restriction of L. pneumophila by a novel mechanism. This work provides a foundation for the use of sorghum as an antimicrobial agent.

microbiology↗