Biology subjects

Warren, E.

Publications and source records attributed to Warren, E..

9 recordsLinked to original sources

bioRxiv2026

Dynamic Exchange of Bacteria and Carbapenem Resistance Genes between Sewer Biofilms and Wastewater

Sewer biofilms represent dynamic interfaces for exchange of bacteria and antibiotic resistance genes between biofilms and the overlying wastewater. Using inline, biofilm reactors, the movement of bacteria and 16S rRNA and carbapenemase genes (blaKPC, blaVIM, blaNDM, blaOXA-48-like, and blaIMP) between wastewater and sewer biofilms was investigated. Established, complex biofilms without these {beta}-lactamase (bla) genes, absorbed resistant bacteria within two minutes of exposure to high concentrations of resistant cultures in lab settings. Carbapenem-resistant organisms from these high-concentration source biofilms transferred to downstream biofilms over 60 minutes of representative sewer shear flows. Mass balances of bacteria and genes in biofilms versus wastewater under representative shear flow showed that biofilms exposed to resistant cultures contributed more to the wastewater than to the downstream biofilms. In field studies, established, complex biofilms without target carbapenem-resistant bacteria and genes from wastewater within hours and then stabilized between 2 to 15 days, not varying by more than 0.5 MPN/cm2 or 0.5 log gene copies (GC)/cm2. In contrast, metagenomic profiles of the bacterial community species continued to change up to 21 days. Established biofilms with resistant bacteria and genes exposed to tertiary-treated wastewater without target carbapenemase genes or meropenem antibiotics did not lose resistant genes or bacteria over nine days of exposure (i.e., < 1 log GC/cm2 reduction). Results show that sewer biofilms contribute to the resistance-gene signal found in sewer wastewater by absorbing and releasing bacteria and genes. Consideration of sewer biofilm dynamics is essential for more accurately interpreting wastewater bacterial concentrations in wastewater-based epidemiology studies. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/726639v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@19f6ce0org.highwire.dtl.DTLVardef@1a507c8org.highwire.dtl.DTLVardef@1a2013dorg.highwire.dtl.DTLVardef@ff8613_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2026

Senescent myoblasts exhibit ROS-dependent Akt-mTORC1 dysregulation and are susceptible to reductive stress-induced cell death.

Ageing is characterised by the accumulation of senescent cells. Owing to their irreversible cell-cycle arrest, these cells lack the capacity to replenish the stem cell pool and regenerate tissue, while their pro-inflammatory secretome propagates senescence in a paracrine manner. Much of the senescent phenotype has been attributed to dysregulated mTORC1 signalling, a key regulator of protein synthesis implicated in organismal ageing. Nonetheless, the mechanism underlying this dysregulation is poorly understood and limited to a few selected cell types. Here, we show that mTORC1 dysregulation is also a characteristic of senescent muscle precursor cells, and in contrast to reports in other cell types, senescent myoblasts do not rely on lysosomal nutrient liberation to sustain mTORC1 activity. Instead, they appear to depend on the PI3K/Akt pathway, which is upregulated in these cells. Exogenous antioxidants were identified to alleviate PI3K/Akt/mTORC1 signalling, while exogenous ROS has the capacity to activate mTORC1, supporting a model in which ROS acts upstream of this pathway in senescent myoblasts. Moreover, antioxidants were able to suppress the expression of pro-inflammatory cytokines and enhance the differentiation of senescent myoblasts. Interestingly, prolonged antioxidant treatment led to increased cell death in senescent but not proliferating myoblasts, suggesting they are more prone to reductive stress-induced cell death. We propose that, in vitro, the antioxidant capacity of many plant-derived compounds may underlie their reported benefits as therapeutics targeting senescent cells (senotherapeutics). Together, our findings provide novel insights into mTORC1-dependent regulation of the senescent phenotype and highlight the role of redox modulation in senotherapeutic strategies.

cell biology↗

bioRxiv2026

AI-Guided CRISPR Screen Accelerates Discovery of New Drug Targets

Psoriasis affects over 125 million people worldwide, yet the mechanistic understanding of keratinocyte-driven inflammation remains incomplete, limiting therapeutic innovation beyond costly systemic biologics that are prone to side effects. Here, we performed the first genome-wide CRISPR knockout screen in primary human adult epidermal keratinocytes to systematically identify regulators of IL-17 receptor A (IL17RA), a central node in psoriatic inflammation. To prioritize therapeutically tractable targets from over 19,000 screened genes, we integrated a large language model - VirtualCRISPR - trained on functional genomics data, identifying arachidonate 5-lipoxygenase (ALOX5) and oxytocin receptor (OXTR) as high-confidence novel hits with minimal prior association with psoriasis. Multi-omics validation revealed that ALOX5 and OXTR regulate IL17RA expression through distinct signaling pathways - ALOX5 through lipid mediators that stabilize the receptor at the cell surface, and OXTR through calcium signaling that reprograms cellular metabolism. Topical delivery of their inhibitors Zileuton (ALOX5) and Cligosiban (OXTR) exhibited therapeutic efficacy comparable to systemic anti-IL17RA antibody in the imiquimod-induced psoriasis model, suppressing pathogenic Th17/Tc17 responses, polarizing macrophages toward anti-inflammatory phenotypes, and normalizing epidermal hyperproliferation. Proteomic profiling in human 3D organotypic skin and murine models confirmed on-target pharmacology and revealed convergent suppression of neutrophil-keratinocyte inflammatory circuits. The use of VirtualCRISPR significantly shortened the timescale from screen to the identification of druggable hits with robust validation, and this work establishes a blueprint for integrating AI-driven target prioritization with functional genomics to accelerate therapeutic discovery.

genomics↗

bioRxiv2025

Evaluation of Detection Methods for Wastewater Surveillance of Antimicrobial-Resistant Bacteria from Healthcare Facilities

Carbapenem resistance is an urgent public health threat. Wastewater surveillance could support antimicrobial resistance monitoring at long-term care facilities (LTCFs). Feasibility of wastewater sampling (via composite or passive sampler, and sewer biofilm swabs) for carbapenemase genes (blaKPC, blaVIM, blaOXA-48-like, blaNDM, and blaIMP) detected by qPCR or GeneXpert(R) Carba-R from a LTCF was assessed over 16 months and compared to clinical infections. blaKPC, blaOXA-48-like, and blaVIM were routinely detected in composite wastewater samples (6.4{+/-}0.8 log10 gene copy (GC)/ml (100% of 61 samples), 5.5{+/-}0.8 (98%), and 5.9{+/-}1.3 (34%), respectively), passive samples (8.2{+/-}0.5 log10 GC/g (31% of 55), 6.6{+/-}0.5 (96%), and 6.6{+/-}0.7 (31%)) and sewer biofilm (6.0{+/-}0.7 log10 GC/cm2 of pipe (100% of 17), 5.0{+/-}0.7 (100%), and 5.8{+/-}2.7 (24%)). Resistomes of wastewater and sewer biofilms differed, but both contained blaKPC, blaVIM, and blaIMP. Passive sampling may be a suitable alternative to composite sampling. Wastewater surveillance is a promising addition to carbapenemase monitoring.

microbiology↗

bioRxiv2025

Ovarian Tumor FAK Inhibition Releases Omega-3 Fatty Acids Stimulating GATA6 Peritoneal Macrophage CXCL13 Production Enhancing Immunotherapy

High grade serous ovarian cancer (HGSOC) is the most lethal gynecologic malignancy in the USA due to chemo- and immuno-therapy resistance. We show that focal adhesion kinase (FAK) inhibition with ifebemtinib or tumor genetic FAK knockout (KO) in syngeneic ovarian tumor models stimulated resident large peritoneal macrophages to express CXCL13 chemokine and promoted B cell infiltration. Macrophage GATA6 inactivation prevented CXCL13 expression and enhanced FAK-KO tumor growth. Combining ifebemtinib with pegylated doxorubicin chemotherapy and anti-TIGIT immune checkpoint antibody extended survival with tumor-associated tertiary lymphoid structure formation. Mechanistically, FAK-KO heat-treated conditioned media contained exosomes enriched with omega-3 fatty acids which stimulated macrophage CXCL13 production. Ifebemtinib-treated tumors, FAK-KO exosomes, and purified eicosapentaenoic acid enhanced murine and human HGSOC-associated tumor macrophage reprogramming and CXCL13 expression. Overall, our studies define a tumor to macrophage signaling linkage via omega-3 exosome lipids supporting B cell recruitment, survival, immunotherapy enhancement, and actionable via small molecule FAK inhibition. eTOC BlurbHigh-grade serous ovarian cancer remains difficult to treat due to therapy resistance. Chen et. al. reveal that tumor FAK inhibition educates macrophages to express CXCL13 associated with B cell infiltration - highlighting a new therapeutic pathway linking FAK inhibition, omega-3 fatty acid containing exosomes, and macrophage mediated anti-tumor activation. Bullet pointsO_LIGenetic or small molecule FAK inhibition enhances ovarian tumor B cell infiltration C_LIO_LITumor FAK inhibition stimulates GATA6+ macrophages to make CXCL13 C_LIO_LIFAKi, pegylated doxorubicin and anti-TIGIT promote tertiary lymphoid structures C_LIO_LIOmega-3 fatty acids stimulate human HGSOC ascites macrophages to make CXCL13 C_LI

cancer biology↗

bioRxiv2025

Endothelial-pericyte interactions regulate angiogenesis via VEGFR2 signaling during retinal development and disease

Pericytes stabilize the microvasculature by enhancing endothelial barrier integrity, resulting in functional networks. During retinal development, pericyte recruitment is crucial for stabilizing nascent angiogenic vasculature. However, in adulthood, disrupted endothelial-pericyte interactions lead to vascular dropout and pathological angiogenesis in ocular microvascular diseases, and strategies to stabilize the retinal vasculature are lacking. We demonstrate that direct endothelial-pericyte contact downregulates pVEGFR2 in endothelial cells, which enhances pericyte migration and promotes endothelial cell barrier function. Intravitreal injection of a VEGFR2 inhibitor in mouse models of the developing retina and oxygen-induced retinopathy increased pericyte recruitment and aided vascular stability. The VEGFR2 inhibitor further rescued ischemic retinopathy by enhancing vascularization and tissue growth while reducing vascular permeability. Our findings offer a druggable target to support the growth of functional and mature microvasculature in ocular microvascular diseases and tissue regeneration overall.

cell biology↗

bioRxiv2025

Derivation of functional retinal endothelial cells from human pluripotent stem cells for therapeutics and modeling

Retinal microvascular diseases involve a compromised inner blood-retina barrier (iBRB), which remains poorly understood. A renewable source of human iBRB endothelium is thus vital for advancing eye research and treatment development. Here, we differentiated human iPSCs into retinal endothelial cells (iRECs) via the Wnt/{beta}-catenin pathway, namely Norrin/Frizzled4 signaling. These iRECs show genetic, protein, and functional fidelity and unique retinal features. When injected into oxygen-induced retinopathy mice, iRECs integrated into the host vascular network and revascularized the ischemic eye, rescuing the tissue. Within microphysiological models, iRECs form perfusable microvascular networks that mimic the iBRBs morphology and phenotype in both health and diabetic retinopathy conditions while also interacting and organizing physiologically with iPSC-derived retinal pericytes. Our studies establish functional human iRECs and microphysiological iBRB models that facilitate mechanistic studies aimed at identifying therapeutic targets and promoting the revascularization of injured retinas, thereby supporting treatment advancement.

bioengineering↗

bioRxiv2024

Positive feedback regulation between RpoS and BosR in the Lyme disease pathogen

In Borrelia burgdorferi, the Lyme disease pathogen, differential gene expression is primarily controlled by the alternative sigma factor RpoS ({sigma}S). Understanding how RpoS levels are regulated is crucial for elucidating how B. burgdorferi is maintained throughout its enzootic cycle. Our recent studies have shown that a homolog of Fur/PerR repressor/activator, BosR, functions as an RNA-binding protein that controls the rpoS mRNA stability. However, the mechanisms of regulation of BosR, particularly in response to host signals and environmental cues, remain largely unclear. In this study, we revealed a positive feedback loop between RpoS and BosR, where RpoS post-transcriptionally regulates BosR levels. Specifically, mutation or deletion of rpoS significantly reduced BosR levels, while artificial induction of rpoS resulted in a dose-dependent increase in BosR levels. Notably, RpoS does not affect bosR mRNA levels but instead modulates the turnover rate of the BosR protein. Furthermore, we demonstrated that environmental cues do not directly influence bosR expression but instead induce rpoS transcription and RpoS production, thereby enhancing BosR protein levels. This discovery adds a new layer of complexity to the RpoN-RpoS pathway and suggests the need to re-evaluate the factors and signals previously believed to regulate RpoS levels through BosR. IMPORTANCELyme disease is the most prevalent arthropod-borne infection in the United States. The etiological agent, Borreliella (or Borrelia) burgdorferi, is maintained in nature through an enzootic cycle involving a tick vector and a mammalian host. RpoS, the master regulator of differential gene expression, plays a crucial role in tick transmission and mammalian infection of B. burgdorferi. This study reveals a positive feedback loop between RpoS and a Fur/PerR homolog. Elucidating this regulatory network is essential for identifying potential therapeutic targets to disrupt B. burgdorferis enzootic cycle. The findings also have broader implications for understanding the regulation of RpoS and Fur/PerR family in other bacteria.

microbiology↗

bioRxiv2023

The great divide: rhamnolipids mediate separation between P. aeruginosa and S. aureus

The coexistence of multiple bacterial species during infection can have significant impacts on pathogenesis. Pseudomonas aeruginosa and Staphylococcus aureus are opportunistic bacterial pathogens that can co-infect hosts and cause serious illness. The factors that dictate whether one species will outcompete the other or whether the two species can coexist are not fully understood. We investigated the role of surfactants in the interactions between these two species on a surface that enables P. aeruginosa to swarm. We found that P. aeruginosa swarms are repelled by colonies of clinical S. aureus isolates, creating physical separation between the two strains. This effect was abolished in mutants of S. aureus that were defective in the production of phenol-soluble modulins (PSMs), which form amyloid fibrils around wild-type colonies. We investigated the mechanism that establishes physical separation between the two species using the Imaging of Reflected Illuminated Structures (IRIS) method, which tracks the flow of the rhamnolipid surfactant layer produced by P. aeruginosa. We found that PSMs produced by S. aureus deflected the rhamnolipid surfactant layer flow, which in turn, altered the direction of P. aeruginosa swarms. These findings show that rhamnolipids mediate physical separation between P. aeruginosa and S. aureus, which enables these species to coexist in distinct microenvironments. Additionally, we found that a Bacillus subtilis surfactant and abiotic hydrophobic molecules repelled P. aeruginosa swarms through surfactant deflection. Our results suggest that surfactant interactions could have major impacts on bacteria-bacteria and bacteria-host relationships. In addition, our findings uncover a mechanism responsible for P. aeruginosa swarm development that does not rely on sensing but instead is guided largely by the flow of the surfactant layer and its boundaries.

microbiology↗