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Robinson, A.

Publications and source records attributed to Robinson, A..

At least 19 recordsLinked to original sources
bioRxiv2026

BLTP3A dysfunction unleashes lysosomal stress-induced antitumor immunity through STING

High-grade serous ovarian cancer progresses within a lipid-rich ascites microenvironment that engages stress-adaptation programs supporting malignant growth, yet how failed stress resolution influences tumor progression remains incompletely defined. Here, we show that unresolved lysosomal stress restrains tumor progression by sustaining tumor cell-intrinsic STING activity and reorganizing antitumor immunity. We identify BLTP3A as a lysosomal stress-resolution factor in ovarian cancer and show that the common germline missense variant BLTP3A M1098T increases stress-induced lysosomal damage. Patients carrying M1098T had longer overall survival and tumors enriched for cytotoxic T cell-myeloid neighborhoods. In syngeneic tumors, Bltp3a loss or orthologous variant expression delayed progression and generated Cxcl9+ antigen-presenting macrophage niches; M1098T-expressing human tumors similarly promoted interferon-responsive myeloid and T-cell enrichment in humanized mice. Mechanistically, ascites-induced lysosomal stress activated a STING-ATF4-BLTP3A feedback program. BLTP3A dysfunction disrupted CASM-associated lysosomal stress resolution, resulting in prolonged STING activity and sustained tumor-derived IFN-{lambda} without increased cGAMP. Tumor-cell STING and IFN-{lambda} were required for immune remodeling and tumor control. Thus, BLTP3A-dependent lysosomal stress resolution constrains the persistence of stress-induced STING signaling, whereas failure of this response sustains a tumor-intrinsic inflammatory program that organizes protective antitumor immunity.

cancer biology↗
bioRxiv2026

Discovery of a molecular glue inhibitor that stabilises a non-productive Kalirin-Rac1 complex

Rho guanosine triphosphatases (GTPases) are molecular-switches implicated in neurodegenerative diseases, yet targeting them through competitive inhibition remains a challenge due to their high affinity for guanine nucleotides. Guanine nucleotide exchange factors (GEFs) catalyse GDP-to-GTP nucleotide exchange to activate GTPases, providing an alternative opportunity for GTPase modulation. Herein, we describe a complex-targeted strategy to inhibit nucleotide exchange with covalent molecular glues that engage the Kalirin-Rac1 GEF-GTPase complex at the nucleotide binding site are sequester the GEF Kalirin. Fragment hits were identified through XChem and in silico screening, and a fragment merging approach resulted in the generation of covalent inhibitors RS-009 and MC-278. Multiple analyses demonstrate our compounds inhibit nucleotide exchange both through competition with the nucleotides and by stabilising a ternary inhibitor-Rac1-Kalirin complex, thereby trapping Kalirin in a non-productive state and reducing GEF turnover. Biochemical selectivity screening and cellular activity-based protein profiling (ABPP) show that selectivity can be achieved across distinct GEF-GTPase complexes, which may result in improved spatiotemporal control over targeting the GTPase alone. This work provides evidence for targeting GTPase signalling via stabilization of the GEF-GTPase complex in a unique covalent molecular glue mechanism and provides the basis of a chemical probe or therapeutic.

biochemistry↗
bioRxiv2026

Recombinant Echis disintegrins reveal distinct inhibitory profiles on platelet aggregation and endothelial cell migration

Disintegrins are snake venom toxins that bind integrins and modulate platelet aggregation and cell migration. Isolation from venom often yields insufficient quantities for comprehensive study, making recombinant expression essential for detailed toxin characterisation. Here, we produced and characterised four disintegrins from Echis coloratus and Echis ocellatus in Escherichia coli. These homologous proteins contain distinct integrin-binding motifs: EcDis_RGD, EoDis_RGD, EcDis_KGD, and EcDis_VGD. Functional activities were evaluated using human platelet aggregation and endothelial cell scratch wound-healing assays. RGD-containing disintegrins exhibited the strongest inhibition of platelet aggregation, with EcDis_RGD showing the highest potency. The EcDis_KGD disintegrin inhibited platelet aggregation less potently, whereas EcDis_VGD showed no detectable activity in our assays. In endothelial cells, EcDis_RGD disintegrin markedly impaired cell migration, while EoDis_RGD displayed weaker anti-migratory activity. Neither KGD- nor VGD-containing disintegrins significantly affected wound closure. EoDis_RGD exhibited similar antiplatelet activity to the native, venom-purified disintegrin ocellatusin, validating our recombinant expression strategy. However, the corresponding PII-SVMP preparation displayed reduced inhibitory activity, consistent with incomplete generation of fully mature disintegrins species. Our findings demonstrate the importance of both integrin-binding motif identity and surrounding sequence context in determining disintegrin function, and support further use of recombinant toxins in toxinology and as therapeutic lead molecules.

biochemistry↗
bioRxiv2026

Disintegrin-like and Cysteine-rich Domains Govern Enzymatic Activity and Substrate Recognition in Echis Snake Venom Metalloproteinases

Snake venom metalloproteinases (SVMPs) are major drivers of pathology following viper envenomation and represent important targets for the development of next-generation recombinant antivenoms. PIII SVMPs are among the most potent haemorrhagic toxins and contain disintegrin-like (Dis) and cysteine-rich (C-rich) accessory domains. Despite their biomedical importance, the mechanistic roles of these accessory domains in substrate recognition and catalysis remain poorly understood. We produced recombinant full-length and domain-deletion variants of two functionally distinct PIII SVMPs: the broadly proteolytic, cytotoxic cPIII and the highly specific prothrombin activator Ecarin. Proteins were expressed as latent zymogens in insect cells, auto-activated by Zn2+, and analysed using enzymatic, blood clotting, and cell-based assays. Progressive removal of the C-rich and Dis domains reduced zymogen auto-activation and markedly diminished catalytic activity in both toxins. In cPIII, domain deletion caused a stepwise loss of proteolytic and cytotoxic activity without altering substrate preference. In Ecarin, removal of the accessory domains strongly impaired prothrombin activation, and thus plasma clotting, demonstrating a critical role in substrate recognition. Conversely, deletion of the C-rich domain increased fibrinogenolytic activity, revealing a substrate-dependent gatekeeping function. Deglycosylation showed that N-linked glycans modulate SVMP activity in a construct-dependent manner. Recombinant Ecarin closely recapitulated the biochemical properties of the native venom-derived toxin. Our data support a model in which PIII SVMP accessory domains enhance substrate positioning and catalytic efficiency while selectively restricting access to non-cognate substrates. These findings establish accessory-domain-mediated substrate recognition as a key determinant of SVMP function, informing rational antivenom design.

biochemistry↗
bioRxiv2026

Off-the-Shelf Multilayer Vascular Grafts with Damage-Resistant Hydrogel Coatings Incorporating Integrin Targeting

Synthetic grafts remain ineffective for small-caliber vascular applications due to thrombosis and intimal hyperplasia. To address these limitations, our lab designed a multilayer graft consisting of a hydrogel coating that promotes post-implantation endothelialization and an electrospun mesh that matches arterial mechanical properties. Damage-resistant hydrogels were engineered using a double-network system composed of polyether urethane diacrylamide and N-acryloyl glycinamide to enhance fracture resistance through hydrogen bonding. In this study, we utilized redox initiation to apply conformal, durable hydrogels to electrospun grafts. Bioactivity wa introduced using streptococcal collagen-like proteins containing 1{beta}1 and 2{beta}1 integrin-binding motifs, enabling selective cell-material interactions that support endothelialization while preserving acute thromboresistance. To establish the feasibility of these grafts as off-the-shelf devices, we evaluated coating integrity and bioactivity retention following sterilization and dynamic physiological loading. Sterilized composites exhibited surgically-associated damage resistance, indicating that sterilization did not compromise hydrogel durability. Coating integrity and bioactivity were also preserved after six weeks of physiological loading. Acut thromboresistance was supported by both static platelet adhesion assays and dynamic whole-blood bioreactor studies using heparinized blood, with low platelet adhesion observed relative to ePTFE. Finally, a pilot ovine carotid model demonstrated successful surgical handling and sustained graft patency. Collectively, these results highlight the promise of multilayer vascular grafts as durable, thromboresistant conduits for small-diameter vascular applications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=60 SRC="FIGDIR/small/741222v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@14ec910org.highwire.dtl.DTLVardef@17259e8org.highwire.dtl.DTLVardef@6cb611org.highwire.dtl.DTLVardef@1251d7d_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗
bioRxiv2026

Cultivation of halophilic archaea in shallow subsurface martian conditions has implications for extant life on Mars

The present-day martian surface is generally considered inhospitable to life because of low atmospheric pressure, intense surface radiation, global desiccation, and oxidizing chemistry which has been increasing since the late Noachian. However, shallow martian subsurface regions where mineralogy has shown groundwater movement may include localized hypersaline environments capable of retaining liquid water and supporting microbial metabolism. Haloferax volcanii, a model halophilic archaeon, has previously been shown to survive under low-pressure martian conditions (24 mbar) and to grow anaerobically supported by the Mars-relevant oxyanions nitrate and perchlorate under high-salinity conditions. Here, we investigated whether H. volcanii could actively grow under a combination of environmental and chemical conditions relevant to potentially habitable shallow subsurface martian lacustrine settings. Cultures were incubated for 160 days under anoxic, CO2-enriched, low-pressure conditions (24 mbar) in hypersaline liquid media supplemented with nitrate or perchlorate. Growth was observed in all low-pressure treatments and was confirmed by increases in optical density and biological reduction of nitrate and perchlorate. Scanning electron microscopy revealed extensive biofilm formation in low-pressure cultures, and Raman spectroscopy demonstrated the persistence of carotenoid biosignatures after prolonged incubation under martian conditions. Water loss remained below 4% across all treatments, indicating long-term stability of hypersaline brines throughout the experiment. These results demonstrate for the first time that a halophilic archaeon is capable of active growth and metabolism under a Mars-relevant combination of low pressure, high salinity, anoxia, and oxidizing chemistry, providing experimental support for the potential habitability of localized shallow subsurface martian environments. ImportanceThe search for cellular life is a major objective of future Mars exploration. While many studies have examined whether microorganisms can survive under martian conditions, far fewer have demonstrated active growth and metabolism. Here, we document Haloferax volcanii as the first halophilic archaeon capable of active growth under a defined combination of Mars-relevant low atmospheric pressure, high salinity, anoxia, and oxidizing chemical conditions. These findings expand the current understanding of the environmental limits of microbial growth and provide experimental evidence that localized brine environments in the shallow martian subsurface could support active microbial metabolism, if suitable organics and liquid water are present. In addition, this study establishes a practical framework for cultivating halophilic microorganisms under low-pressure martian conditions and may help guide future efforts to detect, cultivate, and characterize potential extant life on Mars.

microbiology↗
bioRxiv2026

PRISM : Peptide-specificity annotation of T-cell receptors with uncertainty quantification

Mapping T-cell receptor (TCR) sequences to their cognate peptide-major histocompatibility complex (pMHC) ligands underlies both basic immunology and T-cell target discovery, yet current models aimed at predicting TCR specificity are limited by sparse labels, viral-biased training data, and an inability to recognize receptors outside their training distribution. We present PRISM, an uncertainty-aware metric-learning framework for TCR{beta} sequence representation. PRISM embeds receptors into a peptide-organized latent space, returns top-k peptides by nearest-neighbor retrieval, and abstains on out-of-distribution receptors by modeling an intrinsic uncertainty that tracks annotation correctness. To offset the viral bias of public databases, PRISM augments training data with structure-guided synthetic receptors that diversify TCR sequences while preserving the energetics of the TCR-pMHC interface. Across a held-out set of 923 peptides and the independent IMMREP23 benchmark, PRISM matches or exceeds sequence-based models, with largest gains on rare epitopes. Finally, PRISM learns attention weights on TCR residues that concentrate on the CDR3{beta} salt-bridge and hydrophobic contacts central to peptide recognition, linking PRISMs positional focus to the biochemical properties of TCR-pMHC structures.

immunology↗
bioRxiv2026

Increasing absolute prey community density protects aposematic models and their imperfect Batesian mimics: Evidence from Neotropical Adelpha butterflies

Batesian mimicry is a defensive adaptation where predators learn to avoid aposematic prey and generalize their warning signals to phenotypically similar mimics. The phenotypic accuracy needed for mimics to benefit from this adaptation depends on the relative densities of models and mimics and the models unpalatability. As aposematic models become more unpalatable or more common relative to their mimics, warning signals become stronger, allowing even poor mimics to benefit. However, few studies have disentangled the importance of relative frequencies of models and mimics from absolute density of the prey community (both models and mimics) in driving relaxed selection on imperfect mimics. Here, we test the hypothesis that increasing model unpalatability and absolute prey community density accelerates predator avoidance learning and enhances protection for imperfect mimics. Using replicas of the model Adelpha iphiclus (Linnaeus), its imperfect mimic Adelpha serpa (Boisduval), and the palatable control Junonia evarete (Cramer), we conducted field experiments that enhanced model unpalatability and doubled absolute prey density while maintaining a constant ratio of model, mimic, and control phenotypes. We found that enhanced model unpalatability and increased absolute density significantly reduced predation on all species, highlighting absolute community density as an underappreciated mechanism shaping selection on imperfect Batesian mimics.

ecology↗
bioRxiv2025

The pKa values of buried ionizable amino acids can be determined by the thermodynamic stability of the protein

Ionizable groups in hydrophobic environments in proteins usually titrate with anomalous pKa values. The ionization of these buried residues is often coupled to transitions between conformational states. Here we test the hypothesis that because the thermodynamic stability of a protein ({Delta}G{degrees}H2O) determines the probability of conformational transitions, the apparent pKa values of buried residues can be governed by {Delta}G{degrees}H2O. Variants of staphylococcal nuclease (SNase) with either Lys-66 or Lys-92 buried in its hydrophobic interior were engineered along with surface mutations that alter {Delta}G{degrees}H2O without affecting the electrostatic properties of the internal microenvironments of the buried Lys residues. The measured pKa values of these Lys residues largely correlates with {Delta}G{degrees}H2O. NMR spectroscopy was used to demonstrate that the structural changes of the protein backbone coupled to the ionization of Lys-66 or Lys-92 are comparable regardless of the {Delta}G{degrees}H2O of the protein. NMR spectroscopy confirmed that global unfolding of the Lys-92 variant coincides with the apparent pKa of the Lys side chain. The data presented show that the anomalous pKa values measured for internal residues in proteins do not necessarily report on local dielectric or electrostatic properties of the microenvironments around the ionizable group; rather, they can report on the energetics of pH-driven conformational transitions. These data suggest that accurate structure-based calculation of pKa values will require de novo prediction of partially unfolded conformations, and accurate calculation of free energy differences between conformational states, both of which remain formidable challenges.

biophysics↗
bioRxiv2025

Modeling Hepatocellular Carcinoma and its microenvironment on a chip

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. Its incidence is increasing and is closely related to advanced liver disease. Interactions in the HCC microenvironment between tumor cells and the associated stroma actively regulate tumor initiation, progression, metastasis, and therapy response. Effective drug development increasingly requires advanced models that can be utilized in the earliest stages of compound and target discovery. Here we report a phenotypic screen on an advanced HCC patient-derived chip (PDChip) model. The vascularized HCC PDChip models include relevant cellular players of the HCC microenvironment. We assessed the effect of 28 treatment conditions on a panel of 8 primary HCC tumors and 2 cell lines. Approximately 1200 HCC PDchips were grown under perfusion flow, exposed to treatments, and subsequently assessed for viability, tumor-associated vasculature responses and chemokine and cytokine changes. Although the SoC therapeutics sorafenib and lenvatinib reduced culture viability and produced profound changes in the organization of the vascular beds, they did not affect the tumor cell population in these cultures. Atorvastatin, a 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor, reduced tumor viability but did not affect vascular bed organization. Sorafenib, lenvatinib and atorvastatin also affected chemokine and cytokine release. Tocilizumab, galunisertib, and vactosertib decreased the level of IL6, a relevant prognostic marker for HCC, while IL6 was increased by halofuginone. In conclusion, HCC PDChip models enabled a detailed evaluation of drug-induced responses in the tumor and associated microenvironment, highlighting their importance in preclinical research for understanding diseases and developing new drugs.

bioengineering↗
bioRxiv2025

Advanced Manufacturing of Coil-Reinforced Multilayer Vascular Grafts to Optimize Biomechanical Performance

Small diameter vascular grafts require a complex balance of biomechanical properties to achieve target burst pressure, arterial compliance-matching, and kink resistance to prevent failure. Iterative design of our multilayer vascular was previously used to achieve high compliance while retaining the requisite burst pressure and suture retention strength for clinical use. To impart kink resistance, a custom 3D solution printer was used to add a polymeric coil to the electrospun polyurethane graft to support the graft during bending. The addition of this reinforcing coil increased kink resistance but reduced compliance. A matrix of grafts were fabricated and tested to establish key structure-property relationships between coil parameters (spacing, diameter, modulus) and biomechanical properties (compliance, kink radius). A successful graft design was identified with a compliance similar to saphenous vein grafts (4.1 {+/-} 0.4 %/mmHgx10-2) while maintaining comparable kink resistance to grafts used currently in the clinic. To explore graft combinations that could increase graft compliance to match arterial values while retaining this kink resistance, we utilized finite element (FE) models of compliance and kink radius that simulated experimental testing. The FE-predicted graft compliance agreed well with experimental values. Although the kink model over-predicted the experimental kink radius values, key trends between graft parameters and kink resistance were reproduced. As an initial proof-of-concept, the validated models were then utilized to parse through a targeted graft design space. Although this initial parameter range tested did not yield a graft that improved upon the previous balance of graft properties, this combination of advanced manufacturing and computational framework paves the way for future model-driven design to further optimize graft performance. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=97 SRC="FIGDIR/small/633374v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@faca56org.highwire.dtl.DTLVardef@f9dccforg.highwire.dtl.DTLVardef@fbb5corg.highwire.dtl.DTLVardef@d5c021_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗
bioRxiv2024

Forecasting trade and biosecurity risk under climate change

1.Executive summaryIt is well known that the increasing globalisation of human movement and trade has led to substantial range expansions of many taxa, and as a consequence, is increasingly exposing countries to novel biosecurity threats. This increased exposure to novel threats is expected to be exacerbated by a changing climate as trade patterns and species distributions change. In many countries biosecurity regulators are already struggling to contend with the increased exposure to pests and diseases, and as such, must shift towards a pro-active strategy of anticipating risk. However, to anticipate risk, regulators must make better use of the data they collect (e.g. border interceptions) and develop models capable of forecasting biosecurity propagule pressure and establishment exposure as a function of changing trade, human population, and species distributions. In this project, we develop a highly innovative model framework that simulates annual changes in international trade patterns (imports & exports) as a function of climate change impacts on crop productivity, labour productivity (via heat stress) and the amount of arable land. These simulated changes in global trade are then coupled with border interception data, and annual predicted changes in the geographic distributions of both human populations and threat climate suitability to estimate climate-induced changes in: 1) contamination rates for imports from different trading partners, 2) the total amount of biosecurity risk material arriving at Australias borders, and 3) its exposure to an establishment event (Figure 1.1). O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=91 SRC="FIGDIR/small/601437v1_fig1_1.gif" ALT="Figure 11"> View larger version (32K): org.highwire.dtl.DTLVardef@1755bcforg.highwire.dtl.DTLVardef@1de8225org.highwire.dtl.DTLVardef@1324d1forg.highwire.dtl.DTLVardef@16530e3_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.1.:C_FLOATNO Conceptual diagram of innovative biosecurity forecasting model. The model is made up of two primary sub-modules: 1) The Economic submodel, which incorporates climate change damages (e.g. crop productivity & heat stress) and simulates these impacts on global import and export trade patterns; 2) The Ecological submodel that estimates threat climate suitability, country-by-commodity contamination rates, and then links these to predicted changes in both global trade flows (i.e. outputs from the Economic submodel) and human population to approximate country-specific propagule pressure and establishment exposure. C_FIG Here, we provide an in-depth description of both the trade and biosecurity risk sub-models. We then illustrate the models implementation and outputs under two different climate scenarios (RCP 4.5 and RCP 8.5) for the purposes of forecasting temporal changes in: 1) global and regional economic trade; and 2) the amount of biosecurity risk material arriving at Australias border (i.e. propagule pressure) and what it may mean for establishment exposure. In illustrating the biosecurity risk, we focused on five plant pests deemed by the Department of Agriculture, Fisheries and Forestry (DAFF) as an exemplar of different hitch-hiking functional groups. The exemplar threats examined were: O_LIOverwintering - brown marmorated stink bug (BMSB; Halyomorpha halys) C_LIO_LIEgg laying - Spongy moth (Lymantria dispar) C_LIO_LINesting - Asian honey bee (Apis cerana) C_LIO_LISheltering - Giant African snail (Lissachatina fulica) C_LIO_LIInternal storage - Khapra beetle (Trogoderma granarium). C_LI 2. Key findings & model utility2.1. Global and regional economic trade consequences of climate changeThe climate-enhanced trade model shows significant changes in trade patterns globally. Climate change impacts various countries differently, causing substantial changes in national incomes, productive capacity, and imports and exports around the world. This occurs even with relatively limited damage functions from global warming. Indeed, with limited damage functions (crop productivity losses, labour productivity losses due to heat stress, and loss of arable land from ocean inundation), the impact on national income in Australia from climate change is relatively small (e.g., damages from more substantive sea level rise and storm surge are not included). However, damages to trading partners and changes in imports to Australia, as well as the rest of the world, are considerable. To allow for easy interrogation of these predicted trade impacts, we have developed an interactive dashboard1 that allows users to examine changes in trade patterns - by country, commodity and climate change scenario - for user selected imports by country and total values. The following summarise the key findings from the select damage functions employed within this analysis: O_LIWestern African regions fare the worst in terms of overall GDP impacts, with the region at large experiencing roughly a 20% loss to GDP. The second most affected region is South Asia2 with Pakistan bearing the greatest damages. Some countries emerged relatively unscathed, prominently those located in North America, Europe, Russia, and Oceania. C_LIO_LIIn terms of climate change related damages to key Australian trading partners, we find that losses in China and Southeast Asia3 are the most consequential. Here labour productivity shocks due to heat stress lead to losses in manufacturing sector imports such as Motor vehicles and Electronic equipment. As the largest exporter of manufacturing goods and a key region affected by climate change, China sees the largest decline in exports, however most of South and Southeast Asia fares worse in terms of relative losses with India and Indonesia seeing over 10% losses. Conversely, net gains are made across several European regions, particularly Germany and the UK. C_LIO_LIThe greatest heterogeneity within macroeconomic sector was observed among agricultural imports and in particular crop sectors in large part due to the varied mix of exporters for each crop type and differential impacts experienced. The largest absolute increase among food- and agriculture-related sectors is seen within Vegetables, fruit, nuts with large increases in exports from OECD regions and China. C_LIO_LIThe climate change damage functions employed here likely vastly underestimate the true costs of climate change impacts for a number of reasons. These include the following: O_LIRecent scholarship suggests that large damages to GDP are already being experienced (Romanello et al., 2023) whereas the earlier work used here (with the exception of crop productivity) includes minimal damages at early timesteps. Further, the anticipated impacts from warming may be far larger (particularly at later timesteps) if climate sensitivities have been misestimated (leading to more extreme associated temperature paths) or tipping points are triggered. C_LIO_LIThe application of damages used within the current study is limited in terms of scope, with damages due to human health impacts and changes in tourism trends missing. In addition to the limited scope, some damages that have been implemented have not been fully allocated to all affected sectors. For example, sea level rise is understood entirely within the purview of loss of arable land while physical capital depreciation and impacts to manufacturing hubs located on coastal areas is not currently simulated. C_LIO_LISociopolitical consequences of climate change such as armed conflict and forced migration are not included. Diminished social resilience due to climate change related impacts (e.g., rising food insecurity) has the potential to feed back into the economy in myriad ways, potentially cascading into additional economic damages and/or altered trade patterns. C_LI C_LIO_LIAlthough limited climate change damages have been implemented, large changes in socioeconomic trends including economic and demographic growth are anticipated to occur within the long-term time horizon used within the current study. Such changes affect key Australian trade partners and have the potential to alter the composition of imports at a magnitude equal to or surpassing climate change impacts. Full simulation of both socioeconomic pathways as well as climate change impacts is currently a CEBRA priority. C_LI 2.2. Changes in pest propagule pressure & establishment exposure hitting Australia.While this report focuses on long-term trends in annual propagule pressure and establishment exposure hitting Australias border, the model is capable of estimating such trends for all 70 global regions and can be readily updated with new interception data. As such, to facilitate interrogation of model outputs, we have developed leaflet interactive maps for each of the five threats. Included in these maps are information4 about temporal trends in pest pressure, establishment exposure (if threat is influenced by climate suitability) and proportion of risk attributable to each infected exporters and commodity sector. The following summarise the key findings we found by running the model for the five exemplar threats: O_LIOut of the five exemplar threats, brown marmorated stink bug (BMSB; Halyomorpha halys) posed the greatest propagule pressure and established risk to Australia. This was followed by giant African snail (Lissachatina fulica), Khapra beetle (Trogoderma granarium), Asian honey bee (Apis cerana), and then spongy moth (Lymantria dispar). C_LIO_LIImports of manufacturing commodities (e.g. electronic equipment, motor vehicles and parts, plastics and metals) from infected countries posed the greatest risk of hitch-hiker contamination, with some exporting regions posing greater risk than others. C_LIO_LIWe detected marginal (albeit uncertain) positive impacts of potential area of suitable climate on the contamination rates coming from infected exporting regions for BMSB and to a lesser extent Spongy moth. This meant that in exporting regions where climate suitability is predicted to increase (e.g. Canada & United Kingdom for BMSB), contamination rates will be higher. By contrast, in regions where climate suitability is expected to decrease (e.g. Italy for BMSB), contamination rates are expected to decline. Area of suitable climate had no discernable impacts on contamination rates associated with Asian honey bee or giant African snail.5 C_LIO_LIDifferences in trends in predicted propagule pressure (i.e. number of contaminated lines) hitting Australias border between the two climate scenarios were found to be small and highly uncertain for all threats examined in this report. Multiple reasons are attributable to this: O_LIThere are compensatory effects occurring, whereby reductions (due to either decreased trade or climate-induced declines in contamination likelihoods) in contaminated items from one infected exporter is made up by increases in another. C_LIO_LIClimate impacts on exports and imports of manufactured goods - the commodity types with highest likelihoods of containing most of the modelled hitch-hikers used in this study - are likely to be severely underestimated. Recent work (Romanello et al., 2023) suggests that the existing set of damages included may severely underestimate the true impacts of climate change, especially on manufacturing sectors. Extending the trade model to include a larger set of damage functions, particularly those that encompass physical capital depreciation due to sea level rise and other disasters magnified by climate change (e.g., bushfires), is expected to result in significantly greater economic impacts within affected manufacturing and related sectors. This in turn will manifest in more significant impacts on global export and import flows, and thus, the total pest pressure expected at Australias border. C_LIO_LIThere remains significant uncertainty on how climate suitability in exporting regions impacts contamination rates. While our model detected marginal positive relationships between contamination rates and area of suitable climate for some threats (e.g. BMSB), uncertainty is partly driven by the number of infected exporting countries containing the threat and how variable climate suitability is within and among those trading partners. This uncertainty could be further reduced by including additional interception data from other countries (e.g. New Zealand) as well as incorporating additional information on other factors likely to impact contamination likelihoods such as known extent of established populations, time since establishment, or other factors that can inform population size in exporting country. C_LI C_LIO_LITrends in Australias exposure to potential BMSB or Spongy moth establishment were expected to decline overtime due to a combination of reduced propagule pressure (caused by declines in volume of trade from high risk trading partners and/or climate-induced declines contamination rates), and declines in climatic suitability at Australian locations where goods are likely to be unpacked. These declines were most pronounced under the worst-case climate scenario (RCP 8.5). By contrast, no discernable trends were found for Asian honey bee or giant African snail - mostly due to 1) minor impacts on trade volumes from infected exporters; 2) negligible impacts of climate suitability on contamination rates; and 3) climate suitability in locations import goods are most likely to be unpacked remaining relatively stable (i.e. where urban centres are predicted to remain). C_LI 2.3. Potential model utilityWe believe that this new model provides the necessary information needed for governments and industry to better anticipate - and therefore manage and adapt to - future climate change impacts on domestic and international economies, global trade, and exposure to biosecurity risk. The current work may serve as a foundation for informing policy decisions as well as a range of downstream models. For example model outputs can be used: O_LIto directly inform both short-term and long-term policies associated with border screening effort among different commodities and exporters, threat prioritisation and developing or updating Import Risk Assessments (IRAs). C_LIO_LIas inputs into other models used by DAFF such as: the post-border pest establishment likelihood model edmaps (Camac et al., 2020, 2021b)6 for informing post-border surveillance & establishment potential, the Risk Return Resource Allocation (RRRA) model, and CEBRAs value model (Dodd et al., 2021) for informing potential damages accrued over time by an exotic threat and the benefits of different mitigation strategies. C_LIO_LIto identify regions and economic sectors most susceptible to changing climate, and thus, provide governments with the necessary foresight to plan and adapt to potential impacts. C_LI

ecology↗
bioRxiv2024

Enterobactin and Salmochelin S4 inhibit the growth of Staphylococcus aureus

There is increasing demand for novel antimicrobial agents to tackle the antimicrobial resistance crisis. Here we report that two Enterobacteriaceae-produced siderophores, enterobactin and salmochelin S4, inhibit the growth of Staphylococcus aureus isolates, including methicillin-resistance S. aureus (MRSA) clinical isolates. The MIC50 for different S. aureus isolates were 2-5 M for salmochelin S4 and 5-10 M for enterobactin. This inhibitory activity was partially repressed by adding Fe+3. These siderophores also inhibited the growth of Enterococcus strains, including vancomycin-resistant enterococci (VRE) clinical isolates, though less effectively than for S. aureus. The growth of various Gram-negative bacteria was barely affected by these siderophores. These results shed new light on the role of enterobactin and salmochelin in bacterial physiology and ecology and have potential for the development of novel strategies to combat the rapid rise of multidrug-resistant bacteria.

microbiology↗
bioRxiv2024

Polycaprolactone-based shape memory foams as self-fitting vaginal stents

There is an urgent critical need for a patient-forward vaginal stent that can prevent debilitating vaginal stenosis that occurs in up to 75% of patients who undergo pelvic radiation treatments and adolescent patients after vaginal reconstruction. To this end, we developed a self-fitting vaginal stent based on a shape-memory polymer (SMP) foam that can assume a secondary, compressed shape for ease of deployment. Upon insertion, the change in temperature and hydration initiates foam expansion to shape fit to the individual patient and restore the lumen of the stent to allow egress of vaginal secretions. To achieve rapid actuation at physiological temperature, we investigated the effect of architecture of two photocurable, polycaprolactone (PCL) macromers. Star-PCL-tetraacrylate displayed reduced melting temperature in the target range as compared to the linear-PCL-diacrylate. Emulsion-templating was then used to fabricate foams from 75:25 water-in-oil (W/O) emulsions that were subsequently annealed to yield high-porosity SMP foams. Upon axial shape memory testing, both foams displayed excellent shape fixity (90%); however, only the PCL star-foams displayed shape recovery ([~]84%) at 37{degrees}C to its permanent shape. A custom mold and curing system was then used to fabricate PCL star- foams into hollow, cylindrical stents. The stent was crimped to its temporary insertion shape (50% reduction in diameter, OD [~] 11 mm) with a custom radial crimper and displayed excellent shape fixity for deployment (> 95%) and shape recovery ([~] 100%). To screen vaginal stents, we developed a custom benchtop pelvic model that simulated vaginal anatomy, temperatures, and pressures with an associated computational model. A hysteroscope was used to visualize stent expansion and deformation via a scope port near the cervix of the benchtop model. A crimped SMP vaginal stent was deployed in the model and expanded to walls of the canal ([~]70% increase in cross-sectional area) in less than 5 minutes after irrigation with warm water. The vaginal stent demonstrated retention of vaginal caliber with less than 1% decrease in cross-sectional area under physiological pressure. Collectively, this work demonstrates the potential for SMP foams as self-fitting vaginal stents to prevent stenosis. Additionally, this work provides new open-source tools for the iterative design of other gynecological devices. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/577474v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@1bc3162org.highwire.dtl.DTLVardef@4e5657org.highwire.dtl.DTLVardef@1069d33org.highwire.dtl.DTLVardef@13aa9da_HPS_FORMAT_FIGEXP M_FIG C_FIG Created in BioRender

bioengineering↗
bioRxiv2024

Critical analysis of polycyclic tetramate macrolactam biosynthetic cluster phylogeny and functional diversity

Polycyclic tetramate macrolactams (PTMs) are bioactive natural products commonly associated with certain actinobacterial and proteobacterial lineages. These molecules have been the subject of numerous structure-activity investigations since the 1970s. New members continue to be pursued in wild and engineered bacterial strains, and advances in PTM biosynthesis suggests their outwardly simplistic biosynthetic gene clusters (BGCs) belie unexpected product complexity. Towards addressing the origins of this complexity and understanding its influence on PTM discovery, we engaged in a combination of bioinformatics to systematically classify PTM BGCs, and PTM-targeted metabolomics to compare the products of select BGC types. By comparing groups of producers and BGC mutants, we exposed knowledge gaps that complicate bioinformatics-driven product predictions. In sum, we provide new insights into the evolution of PTM BGCs while systematically accounting for the PTMs discovered thus far. The combined computational and metabologenomic findings presented here should prove useful for guiding future discovery. IMPORTANCEPolycyclic tetramate macrolactam (PTM) pathways are frequently found within the genomes of biotechnologically-important bacteria, including Streptomyces and Lysobacter spp. Their molecular products are typically bioactive, having substantial agricultural and therapeutic interest. Leveraging bacterial genomics for the discovery of new related molecules is thus desirable, but drawing accurate structural predictions from bioinformatics alone remains challenging. This difficulty stems from a combination of previously underappreciated biosynthetic complexity and remaining knowledge gaps, compounded by a stream of yet-uncharacterized PTM biosynthetic loci gleaned from recently sequenced bacterial genomes. We engaged in the following study to create a useful framework for cataloging historic PTM clusters, identifying new cluster variations, and tracing evolutionary paths for these molecules. Our data suggests new PTM chemistry remains discoverable in nature. However, our metabolomic and mutational analyses emphasize practical limitations to genomics-based discovery by exposing hidden complexity.

microbiology↗
bioRxiv2023

Sequential gentle hydration increases encapsulation in model protocells

Small, spherical vesicles are a widely used chassis for the formation of model protocells and investigating the beginning of compartmentalized evolution. Various methods exist for their preparation, with one of the most common approaches being gentle hydration, where thin layers of lipids are hydrated with aqueous solutions and gently agitated to form vesicles. An important benefit to gentle hydration is that the method produces vesicles without introducing any organic contaminants, such as mineral oil, into the lipid bilayer. However, compared to other methods of liposome formation, gentle hydration is much less efficient at encapsulating aqueous cargo. Improving the encapsulation efficiency of gentle hydration would be of broad use for medicine, biotechnology, and protocell research. Here, we describe a method of sequentially hydrating lipid thin films to increase encapsulation efficiency. We demonstrate that sequential gentle hydration significantly improves encapsulation of water-soluble cargo compared to the traditional method, and that this improved efficiency is dependent on buffer composition. Similarly, we also demonstrate how this method can be used to increase concentrations of oleic acid, a fatty acid commonly used in origins of life research, to improve the formation of vesicles in aqueous buffer.

biochemistry↗
bioRxiv2023

The effects of resistance training on denervated myofibers, senescent cells, and associated protein markers in middle-aged adults

Denervated myofibers and senescent cells are hallmarks of skeletal muscle aging. However, sparse research has examined how resistance training affects these outcomes. We investigated the effects of unilateral leg extensor resistance training on denervated myofibers, senescent cells, and associated protein markers in middle-aged participants (MA, 55{+/-}8 years old, 17 females, 9 males). We obtained vastus lateralis (VL) muscle cross-sectional area (mCSA), VL biopsies, and strength assessments before and after training. Fiber cross-sectional area (fCSA), satellite cells (Pax7+), denervated myofibers (NCAM+), senescent cells (p16+ or p21+), senescence-related proteins, and senescence-associated secretory phenotype (SASP) proteins were analyzed from biopsied muscle. Leg extensor peak torque increased after training (p<0.001), while VL mCSA trended upward (p=0.082). No significant changes were observed for fCSA, NCAM+ myofibers, or senescent (p16+ or p21+) cells, albeit satellite cells increased after training (p=0.037). While >90% satellite cells were not p16+ or p21+, most p16+ and p21+ cells were Pax7+ (>90% on average). Training altered 13/46 proteins related to muscle-nerve communication (all upregulated, p<0.05) and 10/19 proteins related to cellular senescence (9 upregulated, p<0.05). Only 1/17 SASP proteins increased with training (IGFBP-3, p=0.031). In conclusion, resistance training upregulates proteins associated with muscle-nerve communication in MA participants but does not alter NCAM+ myofibers. Moreover, while training increases senescence-related proteins in skeletal muscle, this coincided with an increase in satellite cells but not alterations in senescent cell content or SASP proteins. Hence, we interpret these collective findings as resistance training being an unlikely inducer of cellular senescence in humans.

physiology↗
bioRxiv2023

Hydrogel-Polyurethane Fiber Composites With Enhanced Microarchitectural Control For Heart Valve Replacement

Polymeric heart valves offer the potential to overcome the limited durability of tissue based bioprosthetic valves and the need for anticoagulant therapy of mechanical valve replacement options. However, developing a single-phase material with requisite biological properties and target mechanical properties remains a challenge. In this study, a composite heart valve material was developed where an electrospun mesh provides tunable mechanical properties and a hydrogel coating confers an antifouling surface for thromboresistance. Key biological responses were evaluated in comparison to glutaraldehyde-fixed pericardium. Platelet and bacterial attachment were reduced by 38% and 98%, respectively, as compared to pericardium that demonstrated the antifouling nature of the hydrogel coating. There was also a notable reduction (59%) in the calcification of the composite material as compared to pericardium. A custom 3D printed hydrogel coating setup was developed to make valve composites for device-level hemodynamic testing. Regurgitation fraction (9.6 {+/-} 1.8%) and effective orifice area (1.52 {+/-} 0.34 cm2) met ISO 5840-2:2021 requirements. Additionally, the mean pressure gradient was comparable to current clinical bioprosthetic heart valves demonstrating preliminary efficacy. Although the hemodynamic properties are promising, it is anticipated that the random microarchitecture will result in suboptimal strain fields and peak stresses that may accelerate leaflet fatigue and degeneration. Previous computational work has demonstrated that bioinspired fiber microarchitectures can improve strain homogeneity of valve materials toward improving durability. To this end, we developed advanced electrospinning methodologies to achieve polyurethane fiber microarchitectures that mimic or exceed the physiological ranges of alignment, tortuosity, and curvilinearity present in the native valve. Control of fiber alignment from a random fiber orientation at a normalized orientation index (NOI) 14.2 {+/-} 6.9% to highly aligned fibers at a NOI of 85.1 {+/-} 1.4%. was achieved through increasing mandrel rotational velocity. Fiber tortuosity and curvilinearity in the range of native valve features were introduced through a post-spinning annealing process and fiber collection on a conical mandrel geometry, respectively. Overall, these studies demonstrate the potential of hydrogel-polyurethane fiber composite as a heart valve material. Future studies will utilize the developed advanced electrospinning methodologies in combination with model-directed fabrication toward optimizing durability as a function of fiber microarchitecture.

bioengineering↗