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Murphy, B.

Publications and source records attributed to Murphy, B..

At least 19 recordsLinked to original sources
bioRxiv2026

Apoptotic Cells Promote Tim4+ Macrophage Differentiation Through Direct and Indirect Signaling Pathways

Tissue macrophages are the primary cells responsible for efficiently clearing apoptotic cells, yet how professional efferocyte populations are established remains unclear. Tim4 marks a subset of macrophages across multiple tissues that are highly efferocytic, but the mechanisms governing Tim4 acquisition are poorly understood. Here, we show that monocyte-derived macrophages progressively differentiate into Tim4 macrophages in homeostatic and pathological settings. We identify a cell-autonomous pathway in which efficient apoptotic cell clearance promotes Tim4 acquisition in macrophages, as combined deficiency of CD11b and MerTK impairs both efferocytosis and Tim4 acquisition. Surprisingly, excessive apoptotic cell burden enhanced Tim4 acquisition independently of these receptors. We demonstrate that serial exposure to apoptotic cells induces a type 2 immune response characterized by expansion of IL-4-producing populations and that IL-4R-STAT6 signaling is sufficient and required to drive Tim4 acquisition. These findings support a model in which Tim4 acquisition in macrophages integrates direct signals associated with apoptotic cell clearance and indirect signals generated by apoptotic cell burden. We propose that these complementary mechanisms enable tissues to dynamically adjust the size and function of the professional efferocyte compartment to meet changing demands for apoptotic cell clearance.

immunology↗
bioRxiv2026

Bexobrutideg: A Selective, Catalytic Degrader of Bruton's Tyrosine Kinase Overcomes Inhibitor Resistance and Suppresses Autoantibody-Mediated Disease

Brutons tyrosine kinase (BTK) transduces B-cell receptor (BCR), Toll-like receptor (TLR), and Fc receptor (FcR) signaling, and overactivation of these pathways drives B-cell malignancies and antibody-mediated autoimmune disease. Small molecule inhibitors block the enzymatic functions of BTK, but this inhibition is undermined by resistance mutations, several of which abolish BTKs kinase activity yet promote oncogenic signaling through BTK scaffolding functions. We report the discovery and characterization of bexobrutideg (NX-5948), a heterobifunctional degrader that recruits cereblon (CRBN) to selectively degrade BTK while sparing molecular glue neosubstrates. We demonstrate that bexobrutideg acts catalytically, degrading thousands of copies of BTK per molecule per hour, and this event-driven pharmacology renders it resilient to mutations that confer resistance to both covalent- and noncovalent-inhibitors. Bexobrutideg is orally bioavailable, driving deep and durable BTK degradation across species. Bexobrutideg demonstrates strong efficacy in wild-type and ibrutinib-resistant lymphoma models and robustly suppresses pathway activation in models of autoimmune disease.

cancer biology↗
bioRxiv2026

Menstruation is associated with cyclical granulysin peaks in vaginal secretions despite stable expression by cervicovaginal immune cells

ProblemThe anti-microbial protein granulysin is present in vaginal secretions during the follicular phase of the menstrual cycle but nearly disappears during the luteal phase. The reason for this change is unknown. Method of studyParticipants (n = 23) with regular menstrual cycles collected daily vaginal swabs for granulysin ELISAs. Endocervical cytobrushes, ectocervical biopsies, vaginal biopsies, and PBMC were collected across the cycle to enumerate granulysin-expressing cells by flow cytometry. Cycle phase was determined by daily urinary luteinizing hormone testing and confirmed by serum progesterone levels. ResultsGranulysin levels in secretions were up to 10,000 times higher during menstruation than during the luteal phase (menstruation, median 3,924 pg/mL [IQR 400-17,280]; luteal, median and IQR undetectable [<7.81 pg/mL]). In the endocervical canal, granulysin-expressing cells were much more abundant during menstruation than during the mid-follicular or mid-luteal phases. In contrast, the number of granulysin-expressing cells in the ectocervix and vagina remained stable during the cycle. The most abundant granulysin-expressing cell types in the mucosa were CD8 T cells and NK cells. In a minority of participants, granulysin was consistently detected in luteal-phase swabs; this phenomenon was associated with parity. ConclusionsGranulysin in vaginal secretions is associated with menstruation and concomitant with a spike in granulysin-expressing cells in the endocervical canal. This result explains the much higher granulysin levels in secretions during the follicular than the luteal phase. In contrast, immune cells from ectocervical and vaginal biopsies express granulysin independently of the menstrual cycle, indicating their continuous ability to respond to microbial infection.

immunology↗
bioRxiv2025

Genetic Drivers of Sensitivity or Resistance to RAS(ON) Multi-Selective Inhibitors in NRAS-Mutated Melanoma

Most patients with advanced BRAF or NRAS-driven melanoma receive front-line immunotherapy. However, if immunotherapy fails, BRAF-mutated patients have effective second-line therapies, whereas NRAS-mutated patients lack pathway-targeted options. Recently, RAS(ON) multi-selective inhibitors like RMC-7977, and the investigational agent daraxonrasib, were described that, in partnership with cyclophilin-A (CYPA), inhibit RAS[GTP] signaling. Both compounds demonstrate potent anti-proliferative activity against NRAS-mutated melanoma cell lines and robust anti-tumor activity against preclinical melanoma models. However, in preclinical models, resistance to RMC-7977 monotherapy arose through mutations in Ppia (encoding CYPA) or Map2k1 (encoding MEK1). Moreover, two clinical case studies in patients with NRAS-mutated melanoma treated with daraxonrasib demonstrated clear anti-tumor activity in one patient, but progressive disease in another with co-occurring NRAS and MAP2K1 mutations at baseline. These findings support the potential for daraxonrasib in treatment of patients with NRAS-mutated melanoma, and reveal candidate mechanisms of monotherapy resistance, underscoring the need for combination therapies to improve outcomes. SIGNIFICANCEThere are no pathway-targeted therapies for patients with NRAS-mutated melanoma. Here we demonstrate that direct pharmacological inhibition of RAS[GTP] with RMC-7977 or daraxonrasib (RMC-6236) has profound inhibitory effects in preclinical models of NRAS-mutated melanoma. Furthermore, we identify mechanisms of resistance to RMC-7977 through mutational inactivation of CYPA or mutational activation of MEK1.

cancer biology↗
bioRxiv2025

Nanogram of Arg 1 Delivered by extracellular vesicles restore ARG1 activity in a mouse model of ARG1-D and improve lifespan.

Arginase 1 (ARG1) deficiency (ARG1-D) is a rare genetic disorder due to loss of ARG1, the final enzyme in the urea cycle. ARG1-D hepatocytes are impaired in converting arginine into urea, resulting in elevated peripheral arginine and ammonia, which leads to progressive neurological symptoms. Current therapeutic strategies mainly focus on managing plasma arginine and ammonia level, but long-term outcomes remain poor. While no approved treatment specific for ARG1-D is available in the United States, a recombinant protein based enzyme replacement therapy is available in Europe. Recently, extracellular vesicles (EVs) are emerging as a powerful therapeutic vehicle. By using Capricors StealthX platform, EVs were engineered to express human ARG1 on their surface or encapsulated within. Regardless of its localization on the EV membrane, nanograms of ARG1 carried by EVs were biologically active and able to convert arginine into urea as potent as micrograms of human recombinant ARG1 (rHuArg1). Furthermore, ARG1- encapsulating EVs (STX-Arg1-in) were able to deliver ARG1 intracellularly but not EVs carrying ARG1 on their surface or rHuArg1. STX-Arg1-in EVs were further evaluated in a series of in vivo studies and results showed that STX-Arg1-in EVs were non-toxic and able to restore the arginase activities in the liver of the Arg1-/- mice, which leads to a lowered plasma arginine concentration similar to wildtype mice. Most importantly, Arg1-in expanded the life span of the lethal neonatal Arg1 deficiency mouse model. Taken together, our data suggested StealthX-engineered STX-Arg1-in EVs have a better safety profile due to extreme low dosage and had great potential as a novel enzyme replacement strategy for patients suffering from ARG1-D. Significance statementIntracellular delivery of recombinant protein and improved life span are endpoints of a successful enzyme replacement therapy for the treatment of ARG1-D. Using StealthX platform, a fully functional ARG1 enzyme was engineered to be carried inside of the extracellular vesicles, which allowed intracellular delivery of ARG1 protein in vitro and in vivo, with improvement of life span in a lethal neonatal mouse model of Arg1 deficiency. More importantly, no toxicity was observed, and efficacy was achieved with low dose, setting the base for an improved therapeutic approach.

genetics↗
bioRxiv2025

Chemotherapy Induces an IL1β-dependent Neutrophil Recruitment that Promotes Chemoresistance in Metastatic Ovarian Cancer

High-grade serous carcinoma (HGSC) of the ovary acquires chemoresistance through diverse cancer cell-intrinsic and-extrinsic mechanisms, culminating in treatment-refractory intraperitoneal metastasis. How chemotherapy-induced remodeling of the tumor microenvironment modulates drug sensitivity remains unclear. In this study, we demonstrate that chemotherapy induced IL1{beta}-dependent neutrophil accumulation in tumors, driving chemoresistance in HGSC. Using patient samples, bulk transcriptomic profiling before and after chemotherapy revealed post-treatment upregulation of IL1B, and single-cell RNA sequencing identified myeloid cells as its principal source. In a chemoresistant murine metastatic ovarian cancer model, chemotherapy increased neutrophils and neutrophil extracellular traps (NETs) in omentum tumors; these increases were abrogated in IL1{beta}-deficient mice, with expansion of activated CD8+ T cells and tumor control. Neutrophil depletion in wild-type mice recapitulated the chemosensitive phenotype of IL1{beta}-deficient mice. In vitro, IL1{beta} did not alter cancer cell-intrinsic chemosensitivity, whereas NETs reduced the chemosensitivity of cancer cells. Additionally, the dominant IL1{beta} receptor (IL1R1) was predominantly expressed in tumor-associated fibroblasts in humans and mice. Consistently, IL1R1-deficient mice exhibited chemosensitivity with decreased neutrophil accumulation and increased IFN{gamma}TNFCD8 T cells. We also found that chemotherapy upregulated CXCL2 in patients and that ablating IL1{beta}-IL1R1 axis decreased CXCL2 expression in tumor-associated fibroblasts in mice. Finally, residual human HGSC tumor after chemotherapy showed increased neutrophils and a trend toward more NETs. Collectively, these findings illuminate a paradoxical, cancer cell-extrinsic mechanism in HGSC whereby chemotherapy itself amplifies chemoresistance and suggest that targeting chemotherapy-induced inflammation may help overcome treatment resistance.

immunology↗
bioRxiv2025

Characterization of the BRAF interactome identifies BRAFV600E<=>TP53 interaction in melanoma

Melanoma is a highly aggressive and frequently metastatic cancer with its incidence reported to be on the rise. Although most oncogenic drivers in melanoma converge on activation of the RAS>RAF>MEK>ERK MAPK signaling pathway, not all MAPK-activating mutations are recurrently observed in this disease, suggesting a unique functional role for BRAFV600E, which is present in [~]50% of all melanoma cases. However, the prevalence of BRAFV600E alterations over other known MAPK-promoting oncoproteins raises questions regarding whether BRAFV600E possesses additional functions outside of MAPK pathway activation. Thus, we performed TurboID to differentiate the interactome between wild-type BRAF and BRAFV600E. We identified novel interacting partners of normal vs. BRAFV600E, most strikingly being the tumor suppressor TP53. While TP53 is commonly altered or lost across many malignancies, it is notable that TP53 alterations are rare in melanoma. Our studies suggest that BRAFV600E can interact with and inactivate TP53, thus providing potential mechanistic explanation as to why TP53 inactivation or loss is infrequent in BRAFV600E-driven melanoma.

cancer biology↗
bioRxiv2025

BRAFV600E-Driven Lung Tumorigenesis Requires Ligand-Mediated Activation of ERBB Receptor Signaling

Secretion of ligands of the human epidermal growth factor (EGFR) family of receptors or erythroblastic leukemia viral oncogene family (ERBB1-4) is a feature common to many cancer cells. However, our understanding of the role of autocrine ligands in the aberrant behavior of cancer remains incomplete. Here we demonstrate that, in numerous preclinical models of lung tumorigenesis, BRAFV600E signaling promotes expression of ligands including HB-EGF, TGF, Epi- and Amphiregulin. Moreover, using both genetic or pharmacological approaches, we demonstrate that ligand-mediated activation of EGFR signaling in the tumor cell is required to sustain both early-stage BRAFV600E-driven lung tumorigenesis and supports late-stage BRAFV600E-driven lung cancer maintenance. Unbiased Reverse Phase Protein Analyses (RPPA) analyses, paired with targeted validation, reveals ERBB signaling serves to sustain signaling through the ERK1/2 MAP kinase pathway, through effects on ARAF and CRAF, and on the parallel JUN kinase (JNK) pathway. Furthermore, EGFR is activated in a cohort of BRAF-mutated lung cancer patients both pre- and post-treatment. Finally, we noted significant improvement in the depth and durability of therapeutic responses in preclinical models of BRAFV600E-driven lung cancer by combined inhibition of both BRAFV600E signaling plus pan-ERBB signaling. Collectively, this work provides evidence for an important role for ERBB family signaling in the genesis and maintenance of BRAFV600E-driven lung cancers, and the potential for future therapeutic improvement by rational combination targeting of these pathways. SIGNIFICANCEBRAFT1799A serves as a predictive biomarker for FDA-approved targeted inhibition of BRAFV600E oncoprotein kinase signaling in non-small cell lung cancer (NSCLC). However the occurrence of primary or acquired drug resistance limit the depth and durability of patient responses. Studies described here provide a mechanistic rationale for clinical testing of first-line BRAFV600E inhibition combined with pan-ERBB inhibition to improve the depth and durability of initial patient responses, and delay the emergence of lethal drug resistant disease.

cancer biology↗
bioRxiv2025

High-resolution Staphylococcus profiling reveals intra-species diversity in a single skin niche

The skin microbiome is dominated by a few key genera, among which Staphylococcus is one of the most well-characterised. Recent studies have examined the roles of various Staphylococcus species such as S. epidermidis and S. hominis within broader skin microbial communities. However, these investigations often rely on isolates from multiple individuals and hence, limiting their ability to capture intra-community interactions. In this study, we focused on the axillary microbiome of a single healthy individual to characterise the genetic and functional diversity of resident Staphylococcus isolates. Using a low-cost, high-throughput DNA extraction and long-read whole genome sequencing pipeline, we generated complete genomes for 93 isolates spanning seven genetically distinct lineages across three major skin species. These comprised of one dominant with three additional lineages of S. epidermidis, two of S. hominis and one of S. capitis. Functional and metabolic analyses revealed species- and strain-specific features, suggesting potential metabolic cross-feeding and specialisation within this community, including within strains of S. epidermidis. These findings highlight the metabolic complexity and potential interdependence of staphylococci inhabiting a single skin site and the need for strain-level resolution of the community. The strains form part of the York Skin Microbiome (YSM) collection, a growing open biobank of genetically diverse skin isolates from matched individuals.

microbiology↗
bioRxiv2025

Neuroactive steroids activate membrane progesterone receptors to induce sex specific effects on protein kinase activity

Neuroactive steroids (NAS), which are synthesized in the brain from progesterone, exert potent effects on behavior and are used to treat postpartum depression, yet how these compounds induce sustained modifications in neuronal activity are ill-defined. Here, we examined the efficacy of NAS for membrane progesterone receptors (mPRs) {delta} and {varepsilon}, members of a family of GPCRs for progestins that are expressed in the CNS. NAS increase PKC activity via Gq activation of mPR{delta} with EC50s between 3-11nM. In contrast, they activate Gs via mPR{varepsilon} to potentiate PKA activity with similar potencies. NAS also induced rapid internalization of only mPR{delta}. In the forebrain of female mice, mPR{delta} expression levels were 8-fold higher than males. Consistent with this, activation of PKC by NAS was evident in acute brain slices from female mice. Collectively, our results suggests that NAS may exert sex-specific effects on intracellular signaling in the brain via activation of mPRs.

neuroscience↗
bioRxiv2024

Revealing the diversity of commensal corynebacteria from a single human skin site

Our understanding of the skin microbiome has dramatically improved since the pioneering studies and the improvements in sequencing technologies. Species of the genus Corynebacterium are known to form a major part of the human skin microbiome but most detailed studies have focussed on other similarly prevalent genera like Staphylococcus and Cutibacteria. Prior to this study, there were few complete genomes for skin commensals of the genus Corynebacterium, with only 9 complete genomes available for the most commonly identified species C. tuberculostearicum. In this study we explored the genus Corynebacterium from a single body site by swabbing the axilla/underarm of 4 individuals and using a selective media to enrich for corynebacteria. We then generated whole genome sequencing data of these corynebacteria enriched isolates using long-read sequencing and subsequent bioinformatics analysis to reveal an unparalleled diversity of this genus from a single skin site. Through this approach, we obtained the closed genomes of 215 isolates, 154 derived from a single individual. With this genetic information, we were able to identify 7 different species including species previously not associated to the skin and two novel species provisionally named C. axilliensis and C. jamesii. We used pangenome analysis on 30 genetically distinct isolates spanning the 7 species to identify putative metabolic differences, antimicrobial resistance profiles, novel biosynthetic gene clusters (BGCs), prophages and phage defence systems. Our culture-based Nanopore sequencing approach has dramatically improved our overall knowledge of skin corynebacteria, and uniquely here also providing in-depth analysis from a single skin site, revealing a multitude of differences between the isolates. Here we not only improved our knowledge of axillary corynebacteria but also greatly expanded the publicly available number of cutaneous corynebacterial genomes complementing recent studies seeking to understand the diversity of skin corynebacteria.

microbiology↗
bioRxiv2024

Do rhizobia buffer legumes against climate-mediated microbiome effects?

Climate change is altering both soil microbial communities and the ecological context of plant-microbe interactions. Heat, drought, and their legacies can alter soil microbiomes and potential plant symbionts, but the direct consequences of these microbial changes on plant performance and plant investment in symbiosis remain underexplored. Predicting how soil microbes modulate plant resilience to heat and drought is critical to mitigating the negative effects of climate change on ecosystems and agriculture. In this study, we conducted growth chamber experiments to isolate the microbially-mediated indirect effects of heat and drought on plant performance and symbiosis. In the first experiment, focused on drought, we found that drought and drought-treated microbes, along with their interaction, significantly decreased the biomass of Medicago lupulina plants compared to well-watered microbiomes and conditions. In a second experiment, we then tested how the addition of a well-known microbial mutualist, Sinorhizobium meliloti, affected heat- and drought-treated microbiomes impact on M. lupulina. We found that drought-adapted microbiomes negatively impacted legume performance by increasing mortality and reducing branch number, but that adding rhizobia erased differences in plant response to climate treated soils. In contrast, heat-adapted microbiomes did not differ significantly from control microbiomes in their effects on a legume. Our results suggest microbial legacy effects, mutualist partners, and their interactions are important in mediating plant responses to drought, with some mutualists equalizing plant responses across microbial legacies.

ecology↗
bioRxiv2024

Intraperitoneal activation of myeloid cells clears ascites and reveals IL27-dependent regression of metastatic ovarian cancer

Patients with metastatic ovarian cancer (OvCa) have a 5-year survival rate of less than 30% due to persisting dissemination of chemoresistant cells in the peritoneal fluid and the immunosuppressive microenvironment in the peritoneal cavity. Here, we report that intraperitoneal administration of {beta}-glucan and IFN{gamma} (BI) induced robust tumor regression in clinically relevant models of metastatic OvCa. BI induced tumor regression by controlling fluid tumor burden and activating localized antitumor immunity. {beta}-glucan alone cleared ascites and eliminated fluid tumor cells by inducing intraperitoneal clotting in the fluid and Dectin-1-Syk-dependent NETosis in the omentum. In omentum tumors, BI expanded a novel subset of immunostimulatory IL27+ macrophages and neutralizing IL27 impaired BI efficacy in vivo. Moreover, BI directly induced IL27 secretion in macrophages where single agent treatment did not. Finally, BI extended mouse survival in a chemoresistant model and significantly improved chemotherapy response in a chemo-sensitive model. In summary, we propose a new therapeutic strategy for the treatment of metastatic OvCa.

immunology↗
bioRxiv2023

Evaluation of short-term hair follicle storage conditions for maintenance of RNA integrity.

Hair follicles provide an easily accessible tissue for interrogating gene expression in mammals. RNAlater(R) is a liquid storage solution that stabilises and preserves cellular RNA, eliminating the need to immediately process or freeze tissue specimens. The manufacturer advises storage of samples at 2-8{degrees}C overnight before transfer to -20{degrees}C. This study aimed to evaluate RNA integrity in hair follicle samples collected from horses, stabilized in RNAlater(R), and stored under three short-term storage conditions. Mane hair samples complete with follicles were collected from three horses at a single time point. Approximately 15 hairs were placed in each of three 2 mL tubes containing 0.75ml RNAlater(R) solution. Test group A was stored at 4{degrees}C for 24-h, then decanted and stored at -20{degrees}C. Test groups B and C were stored at 4{degrees}C and 19{degrees}C (room temperature) respectively for 7 days, then decanted and stored at -20{degrees}C. RNA was isolated from all samples and RNA quantity and quality were measured. One-way ANOVA revealed no difference in RNA concentration (A:516 +/-125 ng/ml, B:273+/-93 ng/ml, C:476+/-176 ng/ml;P = 0.4293) or quality (A:9.5 +/-0.19, B:9.8+/-0.09, C:9.2+/-0.35 RIN; P = 0.3193) between the test groups. There were no group differences in mean Cycle Threshold values from qPCR validation assays confirming high-quality template cDNA. The results suggest that storage of hair follicles for one week in RNAlater(R) at cool or room temperature conditions will not compromise RNA integrity and will permit extended transport times from remote sampling locations without the need for freezing.

molecular biology↗
bioRxiv2023

An efficient method for high molecular weight bacterial DNA extraction suitable for shotgun metagenomics from skin swabs

The human skin microbiome represents a variety of complex microbial ecosystems that play a key role in host health. Molecular methods to study these communities have been developed but have been largely limited to low-throughput quantification and short amplicon sequencing, providing limited functional information about the communities present. Shotgun metagenomic sequencing has emerged as a preferred method for microbiome studies as it provides more comprehensive information about the species/strains present in a niche and the genes they encode. However, the relatively low bacterial biomass of skin, in comparison to other areas such as the gut microbiome, makes obtaining sufficient DNA for shotgun metagenomic sequencing challenging. Here we describe an optimised high-throughput method for extraction of high molecular weight DNA suitable for shotgun metagenomic sequencing. We validated the performance of the extraction method, and analysis pipeline on skin swabs collected from both adults and babies. The pipeline effectively characterised the bacterial skin microbiota with a cost and throughput suitable for larger longitudinal sets of samples. Application of this method will allow greater insights into community compositions and functional capabilities of the skin microbiome. Impact StatementDetermining the functional capabilities of microbial communities within different human microbiomes is important to understand their impacts on health. Extraction of sufficient DNA is challenging, especially from low biomass samples, such as skin swabs suitable for shotgun metagenomics, which is needed for taxonomic resolution and functional information. Here we describe an optimised DNA extraction method that produces enough DNA from skin swabs, suitable for shotgun metagenomics, and demonstrate it can be used to effectively characterise the skin microbiota. This method will allow future studies to identify taxonomic and functional changes in the skin microbiota which is needed to develop interventions to improve and maintain skin health. Data SummaryAll sequence data and codes can be accessed at: NCBI Bio Project ID: PRJNA937622 DOI: https://github.com/quadram-institute-bioscience/coronahit_guppy DOI: https://github.com/ilianaserghiou/Serghiou-et-al.-2023-Codes

genomics↗
bioRxiv2021

High-throughput Activity Assay for Screening Inhibitors of the SARS-CoV-2 Mac1 Macrodomain

Macrodomains are a class of conserved ADP-ribosylhydrolases expressed by viruses of pandemic concern, including coronaviruses and alphaviruses. Viral macrodomains are critical for replication and virus-induced pathogenesis; therefore, these enzymes are a promising target for antiviral therapy. However, no potent or selective viral macrodomain inhibitors currently exist, in part due to the lack of a high-throughput assay for this class of enzymes. Here, we developed a high-throughput ADP-ribosylhydrolase assay using the SARS-CoV-2 macrodomain Mac1. We performed a pilot screen which identified dasatinib and dihydralazine as ADP-ribosylhydrolase inhibitors. Importantly, dasatinib does not inhibit MacroD2, the closest Mac1 homolog in humans. Our study demonstrates the feasibility of identifying selective inhibitors based on ADP-ribosylhydrolase activity, paving the way for screening large compound libraries to identify improved macrodomain inhibitors and explore their potential as antiviral therapies for SARS-CoV-2 and future viral threats.

biochemistry↗
bioRxiv2021

AMP-Kinase mediates regulation of glomerular volume and podocyte survival

We reported that Shroom3 knockdown, via Fyn inhibition, induced albuminuria with foot process effacement (FPE) without glomerulosclerosis (FSGS) or podocytopenia. Interestingly, knockdown mice had reduced podocyte volumes. Human minimal change disease, where podocyte Fyn inactivation was reported, also showed lower glomerular volumes than FSGS. We hypothesized that lower glomerular volume prevented the progression to podocytopenia. To test this hypothesis, we utilized unilateral- and 5/6th nephrectomy models in Shroom3 knockdown mice. Knockdown mice exhibited lower glomerular volume, and less glomerular and podocyte hypertrophy after nephrectomy. FYN-knockdown podocytes had similar reductions in podocyte volume, implying Fyn was downstream of Shroom3. Using SHROOM3- or FYN-knockdown, we confirmed reduced podocyte protein content, along with significantly increased phosphorylated AMP-kinase, a negative regulator of anabolism. AMP-Kinase activation resulted from increased cytoplasmic redistribution of LKB1 in podocytes. Inhibition of AMP-Kinase abolished the reduction in glomerular volume and induced podocytopenia in mice with FPE, suggesting a protective role for AMP-Kinase activation. In agreement with this, treatment of glomerular injury models with AMP-Kinase activators restricted glomerular volume, podocytopenia and progression to FSGS. In summary, we demonstrate the important role of AMP-Kinase in glomerular volume regulation and podocyte survival. Our data suggest that AMP-Kinase activation adaptively regulates glomerular volume to prevent podocytopenia in the context of podocyte injury.

systems biology↗
bioRxiv2021

Capturing site-to-site variability through Hierarchical Bayesian calibration of a process-based dynamic vegetation model

Process-based ecosystem models help us understand and predict ecosystem processes, but using them has long involved a difficult choice between performing data- and labor-intensive site-level calibrations or relying on general parameters that may not reflect local conditions. Hierarchical Bayesian (HB) calibration provides a third option that frees modelers from assuming model parameters to be completely generic or completely site-specific and allows a formal distinction between prediction at known calibration sites and "out-of-sample" prediction to new sites. Here, we compare calibrations of a process-based dynamic vegetation model to eddy-covariance data across 12 temperate deciduous Ameriflux sites fit using either site-specific, joint cross-site, or HB approaches. To be able to apply HB to computationally demanding process-based models we introduce a novel emulator-based HB calibration tool, which we make available through the PEcAn community cyberinfrastructure. Using these calibrations to make predictions at held-out tower sites, we show that the joint cross-site calibration is falsely over-confident because it neglects parameter variability across sites and therefore underestimates variance in parameter distributions. By showing which parameters show high site-to-site variability, HB calibration also formally gives us a structure that can detect which process representations are missing from the models and prioritize errors based on the magnitude of the associated uncertainty. For example, in our case-study, we were able to identify large site-to-site variability in the parameters related to the temperature responses of respiration and photosynthesis, associated with a lack of thermal acclimation and adaptation in the model. Moving forward, HB approaches present important new opportunities for statistical modeling of the spatiotemporal variability in modeled parameters and processes that yields both new insights and improved predictions.

ecology↗