Biology subjects

Hansen, J.

Publications and source records attributed to Hansen, J..

At least 19 recordsLinked to original sources

bioRxiv2026

Capturing Spatial Heterogeneity Using UAV-Derived Multispectral Phenotypes and Random Regression Models Improves Genetic Evaluation in Alfalfa Breeding Trials

Spatial heterogeneity is a major source of environmental noise in field trials, reducing genetic evaluation accuracy. Conventional spatial correction methods use field layout and plot proximity but often fail to capture in-season dynamics or leverage temporal data from high-throughput phenotyping. We hypothesized that longitudinal UAV-derived multispectral measurements could characterize field-scale environmental heterogeneity and improve genetic evaluation in breeding trials. Multispectral imagery was collected throughout the growing season across eight alfalfa trials in New York and New Mexico (20222023). Vegetation indices were evaluated for heritability and association with biomass yield using univariate and multi-trait models. Longitudinal measurements were modeled with random regression (RR) to estimate plot-specific permanent environmental effects (PEE), incorporated into genomic mixed models within a two-stage framework. Model performance was compared with conventional autoregressive (AR) and baseline (no correction) models using metrics of consistency and accuracy of estimated genetic effects across replicates. Vegetation indices showed moderate to high heritability, with GNDVI consistently outperforming NDVI and NDRE. Both univariate and multi-trait analyses revealed substantial spatial variability, particularly in New Mexico trials. RR-derived PEE captured field-scale spatial patterns and improved genetic signal recovery relative to baseline and AR models, with the largest gains under strong spatial heterogeneity and comparable performance where variability was limited. The RR framework improved reproducibility of genetic effect estimates across independent field subsets without compromising agreement with full-trial evaluations. These findings show that integrating UAV data via random regression enhances genetic signal recovery and selection accuracy, with broader potential for precision agriculture and biosensor-driven crop management.

genetics↗

bioRxiv2025

Evaluating effects of elevated and accelerated NPQ relaxation on yield, physiology and transcription in soybean

Improving photosynthetic efficiency is proposed as a means of enhancing crop productivity under rising atmospheric CO2 concentrations ([CO2]). To assess whether additive gains can be realized from simultaneous improvements in light use-efficiency and carbon assimilation, we investigated the physiological and molecular responses to elevated CO2 (e[CO2]) of a transgenic soybean line (VPZ-34A) expressing the genes violaxanthin de-epoxidase (VDE), Photosystem II subunit S (PsbS) and zeaxanthin epoxidase (ZEP) from Arabidopsis thaliana (arabidopsis) (AtVPZ). Plants were grown at a free-air CO2 enrichment facility (SoyFACE) in 2021, with physiological and transcriptomic measurements collected during vegetative (V4-5) and reproductive stages (R5-6), then compared to final biomass and seed production parameters. VPZ-34A showed increased maximum quantum photosynthetic efficiency of photosynthesis ({varphi}PSIImax) under fluctuating light during vegetative stages, but no increase in maximum efficiency of carbon assimilation ({varphi}CO2, max), and no significant increase in seed or biomass related traits under ambient or e[CO2]. Transcriptome analysis showed the VPZ construct impacted a small number of genes, among them downregulation of a mitochondrial thioredoxin (Trx o1) and up-regulation of two AP2 transcription factors in VPZ lines may suggest alterations in redox balance and seed development. In conclusion, the improvements in {varphi}PSIImax and NPQ in VPZ-34A were unaffected by e[CO2] but didnt translate to stimulation of yield. Our data highlight the complexity of VPZ manipulation on plant performance and provide avenues for future research to optimize this strategy to improve crop yields.

plant biology↗

bioRxiv2025

Cellular and Spatial Drivers of Unresolved Injury and Functional Decline in the Human Kidney

Building upon a foundational Human Kidney resource, we present a comprehensive multi-modal atlas that defines spatially resolved versus unresolved repair states and mechanisms in human kidney disease. Homeostatic interactions between injured kidney epithelium and its surrounding milieu determine successful repair outcomes, while pathogenic signaling promotes unresolved inflammation and fibrosis leading to chronic disease. We integrated multiple single-cell and spatial modalities across [~]700 samples from >350 patients ([~]250 research biopsies), analyzing [~]1.7 million cells alongside complementary mouse multi-omic profiles spanning acute-to-chronic injury and aging (>300,000 cells) and spatial transcriptomic analysis of >150 human biopsies. This cross-species atlas delineates functional pathways and druggable targets across the nephron and defines gene regulatory networks and chromatin landscapes governing tubular, fibroblast, and immune cell transitions from injury to either recovery or failed repair states. We identified distinct cellular states associated with specific pathological features that show dynamic distributions between acute kidney injury (AKI) and chronic kidney disease (CKD), organized within unique spatial niches that reveal progression mechanisms from early injury to unresolved disease. Gene regulatory analyses prioritized key transcription factor activities (SOX4, SOX9, NFKB1, REL, KLFs) and their target networks establishing disease states and tissue microenvironments. These regulatory programs were directly linked to clinical outcomes, identifying molecular signatures of recovery and secreted biomarkers predictive of AKI-to-CKD progression, providing a key resource for therapeutic development and precision medicine approaches in kidney disease.

molecular biology↗

bioRxiv2025

predicTox: An integrated database of clinical risk frequencies and human gene expression signatures for cardiotoxic drugs

We have used drug-induced transcriptomic responses and whole-genome sequences in healthy human induced pluripotent stem cells (iPSC)-derived cardiomyocyte lines to identify cellular functional pathways and genomic variants potentially associated with the cardiotoxic effects of tyrosine kinase inhibitors (TKIs) and other cancer drugs. Here, we describe predicTox.org, an interactive website that organizes our data and its integration with knowledge from cell pathways and genomic databases. DrugTox summary cards give results of these analyses and metadata for each drug. Fields include FDA Adverse Event Reporting System (FAERS) cardiotoxicity risk scores, cell pathways, and genomic variants potentially associated with drug-induced cardiotoxicity. At a detailed level, predicTox provides ranked lists of pathway signatures that are common for cardiotoxic TKIs, up- and downregulated pathways associated with cardiotoxicity induced by specific TKIs and TKI-regulated genes mapping to those pathways. predicTox provides downloadable lists of drug-induced differentially expressed genes (DEGs) and pathways, drug-related genomic variants associated with cardiotoxicity, all with their statistical metrics, and mathematical models to simulate drug effects on heart physiology. Building on the results of our algorithm for independent reidentification of the well-known rs2229774 variant for anthracycline-induced cardiotoxicity, we describe how our data can be queried to identify potential variants associated with drug-induced cardiotoxicity by affecting a drugs pharmacodynamics and pharmacokinetics.

cancer biology↗

bioRxiv2025

Modulation of the GT Family 47 clade B gene affects arabinan deposition in elaters of Marchantia polymorpha

The plant cell wall polymer -1,5-linked arabinan is associated with important functions in plant physiology, such as cell wall flexibility and desiccation tolerance in angiosperms. However, its biosynthetic mechanism remains obscure. A putative arabinan arabinosyltransferase (AraT) is the Arabidopsis ARABINAN-DEFICIENT1 in GT family 47 clade B (GT47B), but its role is inferred by its mutant cell wall phenotypes, and not through in vitro assays. With seven other genes in the clade, investigating the function of the members in GT47B is hampered by the high genetic redundancy in Arabidopsis. Instead, we probe the function of the only two genes in GT47B in the liverwort model organism Marchantia polymorpha, named MpARAD-Like 1 (MpARADL1) and MpARADL2. Mparadl1 and Mparadl2 loss-of-function mutants and overexpression lines were generated, and their cell wall composition was probed using Comprehensive Microarray Polymer Profiling (CoMPP), glycosyl linkage analysis and immunolabelling. Interestingly, Mparadl2 mutants have much less of the 1,5--L-arabinan epitope in elaters since LM6 antibody recognition showed a notable reduction. However, 5-linked Araf levels in the Marchantia thallus tested with glycosyl linkage analysis are comparable between mutants and wild type, suggesting that there is another enzyme forming 5-Araf linkages in Marchantia. Our attempts to obtain the biochemical activity of these enzymes through the expression and purification of AtARAD1, MpARADL1 and MpARADL2 proteins in the HEK293 cell heterologous expression system were unsuccessful. Therefore, we suspect that an evolutionarily conserved, and clade-specific mechanism is required to confer solubility for purification and subsequent activity characterisation of GT47B proteins. Previous studies have shown that co-expression of protein interaction partners can enhance protein solubility in the HEK293 cell system, but co-expression of MpARADL2 with AtARAD1 and MpARADL1 did not yield soluble proteins, prompting further investigation into the protein-protein interactors of GT47B proteins.

plant biology↗

bioRxiv2025

Functional profiling reveals a non-enzymatic role of NUDT5 in repressing purine de novo synthesis

Folate metabolism is intricately linked to purine de novo synthesis through the incorporation of folate-derived one-carbon units into the purine scaffold. Here, we investigate the chemical and genetic dependencies caused by mutations in the folate enzyme MTHFD1 and discover a key role for Nudix hydrolase 5 (NUDT5) in regulating purine de novo synthesis. Through genetic knockout and development of a selective chemical NUDT5 degrader, we uncover an unprecedented scaffolding role rather than NUDT5 enzymatic activity is responsible for this phenotype. We find that NUDT5 interacts with the rate-limiting enzyme of purine de novo synthesis, PPAT, to repress the pathway in response to elevated purine levels. Our findings establish NUDT5 as an important regulator of purine de novo synthesis and elucidate its role in mediating sensitivities to 6-thioguanine in cancer treatment and to adenosine in MTHFD1 deficiency.

molecular biology↗

bioRxiv2025

MBCO PathNet: Integration and visualization of networks connecting functionally related pathways predicted from transcriptomic and proteomic datasets

Our desktop application MBCO PathNet allows for quick and easy integration and visualization of networks of functionally related pathways predicted from numerous gene and protein lists using the Molecular Biology of the Cell Ontology (MBCO) and other ontologies. Within networks of hierarchical parent-child relationships or functional relationships, pathways are visualized as pie charts where each slice represents a dataset that predicted that pathway. Sizes of pies and slices can be selected to represent statistical significance or other quantitative measures. In addition, MBCO PathNet can generate bar diagrams, heatmaps and timelines. Availability and implementationmbc-ontology.org github.com/SBCNY/Molecular-Biology-ofthe-Cell iyengarlab.org/mbcpathnet Contact: jens.hansen@mssm.edu

bioinformatics↗

bioRxiv2024

Mapping atherogenesis mechanisms in smooth muscle cells by targeting genes linked to coronary artery disease

Recent genome-wide association studies (GWAS) have identified multiple vascular cell-expressed genes linked to coronary artery disease (CAD), suggesting that smooth muscle cells (SMCs) and SMC-derived metaplastic cells are promising targets for novel antiatherosclerosis therapies. However, the disease-promoting pathways of most GWAS-identified genes are unknown, hindering their translation into therapeutic targets. This study integrated public GWAS data for CAD and single-cell RNA sequencing (scRNA-seq) analyses of human atherosclerotic plaques to identify 20 GWAS risk genes with a putative mechanism of action in SMCs or SMC-derived cells. Gene perturbation experiments in SMCs coaxed to plaque-relevant phenotypes revealed that the selected risk genes, despite encoding very different types of proteins, regulated shared sets of genes associated with contractile functions, cell cycle pathways, NF{kappa}B, and type I interferon signaling. By integrating information about GWAS gene effect direction and a deep analysis of cholesterol- and stretch-induced gene modules in SMCs, we find evidence that cholesterol-induced signaling is a pro-atherogenic disease mechanism in SMCs that is upregulated by detrimental and downregulated by protective GWAS genes. Overall, our study identifies a set of candidate disease mechanisms in SMCs that are regulated by multiple GWAS genes across several SMC assays. Furthermore, it provides proof-of-concept for using GWAS gene effect directionality to predict the pathogenic effect of candidate disease mechanisms that can be extended to other GWAS genes and cell types in the future.

cell biology↗

bioRxiv2024

Intrinsic Diversity in Primary Cilia Revealed Through Spatial Proteomics

Primary cilia are a critical organelle found on most human cells, and their dysfunction is linked to hereditary ciliopathies with a wide phenotypic spectrum. Despite their significance, the specific roles of cilia in different cell types remain poorly understood due to limitations in analyzing ciliary protein composition. We employed antibody-based spatial proteomics to expand the Human Protein Atlas to primary cilia. Our analysis identified the subciliary locations of 715 proteins across three cell lines, examining 128,156 individual cilia. We found that 69% of the ciliary proteome is cell-type specific, and 78% exhibited single-cilia heterogeneity. Our findings portray cilia as sensors tuning their proteome to effectively sense the environment and compute cellular responses. We identified 91 novel cilia proteins and found a genetic candidate variant in CREB3 in one clinical case with features overlapping ciliopathy phenotypes. This open, spatial cilia atlas advances research on cilia and ciliopathies.

cell biology↗

bioRxiv2024

Direct β1/β2 AMPK activation reduces liver steatosis but not fibrosis in a mouse model of non-alcoholic steatohepatitis

5AMP-activated protein kinase (AMPK) activators show potential for treating Non-Alcoholic Fatty Liver Disease (NAFLD) and Non-Alcoholic Steatohepatitis (NASH) due to their inhibiting effects on fatty acid and cholesterol synthesis. The absence of treatments for NASH, and its propensity for progression to severe disease, lead us to identify and characterize BI9774, a small molecule AMPK activator, which we used to evaluate this potential, including its ability to reduce the NASH specific qualities of fibrosis and inflammation in a preclinical study. Male Lepob/Lepob mice on a control or NASH inducing (AMLN) diet, with liver fibrosis were given BI9774 or vehicle for 6 weeks while metabolic and NASH endpoints were evaluated. BI9774 treatment decreased plasma ALT, terminal liver weight, and liver lipids. RNA expression of collagen-related genes decreased, although collagen protein and inflammation remained unaltered. We also observed increased heart weight and glycogen levels, and increased expression of genes associated with cardiac hypertrophy. AMPK activation improved many metabolic endpoints, but lack of significant improvement in liver fibrosis and negative cardiac effects suggest systemic AMPK activation is not an ideal NASH therapy. Reductions in steatosis and fibrosis-related genes indicate that, with extended treatment, a liver specific AMPK activator has potential to resolve hepatic fibrosis. Summary StatementFatty liver disease affects up to 30 percent of adults worldwide with 30% of patients progressing to more sever liver disease. AMPK activation can help reduce liver fat.

pharmacology and toxicology↗

bioRxiv2024

Genetic gradual reduction of OGT activity unveils the essential role of O-GlcNAc in the mouse embryo

The reversible glycosylation of nuclear and cytoplasmic proteins (O-GlcNAcylation) is catalyzed by a single enzyme, namely O-GlcNAc transferase (OGT). The mammalian Ogt gene is X-linked and it is essential for embryonic development and for the viability of proliferating cells. We perturbed OGTs function in vivo by creating a murine allelic series of four single amino acid substitutions reducing OGTs catalytic activity to a range of degrees. The severity of the embryonic lethality was proportional to the degree of impairment of OGTs catalysis, demonstrating that the O-GlcNAc modification itself is required for early development. We identified milder hypomorphic Ogt alleles that perturb O-GlcNAc homeostasis while being compatible with embryogenesis. The analysis of the transcriptomes of the mutant embryos at different stages suggested a sexually-dimorphic developmental delay caused by the decrease in O-GlcNAc. Furthermore, a mild reduction of OGTs enzymatic activity was sufficient to loosen the silencing of endogenous retroviruses in vivo.

developmental biology↗

bioRxiv2024

Network Spreading and Local Biological Vulnerability in Amyotrophic Lateral Sclerosis

Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disease that predominantly targets the motor system. Spread of pathology is thought to be driven by both local vulnerability and network architecture. Namely, molecular and cellular features may confer vulnerability to specific neuronal populations, while synaptic contacts may also increase exposure to pathology in connected neuronal populations. However, these principles are typically studied in isolation and it remains unknown how local vulnerability and network spreading interact to shape cortical atrophy. Here we investigate how network structure and local biological features jointly shape the spatial patterning of atrophy in ALS. We analyze the Canadian ALS Neuroimaging Consortium (CALSNIC) dataset and estimate cortical atrophy using deformation-based morphometry (DBM). We find that structural connectivity closely aligns with the course of atrophy. Atrophy is also more likely to occur in regions that share similar transcriptomic, neurotransmitter receptor and metabolic profiles. We identify disease epicenters in motor cortex. Epicenter probability maps show transcriptomic enrichment for biological pathways involved in mitochondrial function as well as support cells, including endothelial cells and pericytes. Finally, individual differences in epicenter location correspond to individual differences in clinical and cognitive symptoms, and differentiate patient subtypes.

neuroscience↗

bioRxiv2024

Remote sensing for estimating genetic parameters of biomass accumulation and modeling stability of growth curves in alfalfa

Multi-spectral imaging (MSI) collection by unoccupied aerial vehicles (UAV) is an important tool to measure growth of forage crops. Information from estimated growth curves can be used to infer harvest biomass and to gain insights in the relationship of growth dynamics and harvest biomass stability across cuttings and years. In this study, we used MSI to evaluate Alfalfa (Medicago sativa L. subsp. sativa) to understand the longitudinal relationship between vegetative indices (VIs) and forage/biomass, as well as evaluation of irrigation treatments and genotype by environment interactions (GEI) of different alfalfa cultivars. Alfalfa is a widely cultivated perennial forage crop grown for high yield, nutritious forage quality for feed rations, tolerance to abiotic stress, and nitrogen fixation properties in crop rotations. The direct relationship between biomass and VIs such as Normalized difference vegetation index (NDVI), green normalized difference vegetation index (GNDVI), red edge normalized difference vegetation index (NDRE), and Near infrared (NIR) provide a non-destructive and high throughput approach to measure biomass accumulation over subsequent alfalfa harvests. In this study, we aimed to estimate the genetic parameters of alfalfa VIs and utilize longitudinal modeling of VIs over growing seasons to identify potential relationships between stability in growth parameters and cultivar stability for alfalfa biomass yield across cuttings and years. We found VIs of GNDVI, NDRE, NDVI, NIR and simple ratios to be moderately heritable with median values for the field trial in Ithaca, NY to be 0.64, 0.56, 0.45, 0.45 and 0.40 respectively, Normal Irrigation (NI) trial in Leyendecker, NM to be 0.3967, 0.3813, 0.3751, 0.3239 and 0.3019 respectively, and Summer Irrigation Termination (SIT) trial in Leyendecker, NM to be of 0.11225, 0.1389, 0.1375, 0.2539 and 0.1343, respectively. Genetic correlations between NDVI and harvest biomass ranged from 0.52 - .99 in 2020 and 0.08 - .99 in 2021 in the NY trial. Genetic correlations for NI trial in NM for NDVI ranged from 0.72 - .98 in 2021 and SIT ranged from 0.34-1.0 in 2021. Genotype by genotype by interaction (GGE) biplots were used to differentiate between stable and unstable cultivars for locations NY and NM, and Random regression modeling approaches were used to estimate growth parameters for each cutting. Results showed high correspondence between stability in growth parameters and stability, or persistency, in harvest biomass across cuttings and years. In NM, the SIT trial showed more variation in growth curves due to stress conditions. The temporal growth curves derived from NDVI, NIR and Simple ratio were found to be the best phenotypic indices on studying the stability of growth parameters across different harvests. The strong correlation between VIs and biomass present opportunities for more efficient screening of cultivars, and the correlation between estimated growth parameters and harvest biomass suggest longitudinal modeling of VIs can provide insights into temporal factors influencing cultivar stability.

genetics↗

bioRxiv2024

BZR1 promotes pluripotency acquisition and callus development through direct regulation of ARF7 and ARF19

Plants have the remarkable ability to regenerate whole organisms through formation of pluripotent cell masses from somatic cells. Cellular programs leading to fate change of somatic to pluripotent cells resembles lateral root (LR) formation and both are chiefly regulated by auxin. Brassinosteroid signalling also plays an important role during LR formation but little is known about the direct link between auxin and brassinosteroid components, such as BZR1 and BES1, in relation to pluripotency acquisition. Here we show that gain-of-function mutants bzr1-D and bes1-D exhibit altered callus formation, yet disruption of these transcription factors does not produce major changes to callus formation or de novo organogenesis. Moreover, our data reveals that BZR1 displays enhanced expression in callus tissue and directly binds to the promoters of ARF7 and ARF19, two master pluripotency regulators, leading to their enhanced transcription. Remarkably, we see abrogation of callus formation in bzr1-D upon disruption of ARF7 and ARF19, emphasizing that BZR1 callus phenotype is dependent on these two auxin signalling components. In conclusion, we depict a link between ARF7, ARF19 and BZR1 in the promotion of pluripotency acquisition, portraying BZR1 as a major supporting factor in callus formation. IMPORTANTO_LIManuscripts submitted to Review Commons are peer reviewed in a journal-agnostic way. C_LIO_LIUpon transfer of the peer reviewed preprint to a journal, the referee reports will be available in full to the handling editor. C_LIO_LIThe identity of the referees will NOT be communicated to the authors unless the reviewers choose to sign their report. C_LIO_LIThe identity of the referee will be confidentially disclosed to any affiliate journals to which the manuscript is transferred. C_LI GUIDELINESO_LIFor reviewers: https://www.reviewcommons.org/reviewers C_LIO_LIFor authors: https://www.reviewcommons.org/authors C_LI CONTACTThe Review Commons office can be contacted directly at: office@reviewcommons.org

plant biology↗

bioRxiv2024

Ontology-based modeling, integration, and analysis of heterogeneous clinical, pathological, and molecular kidney data for precision medicine

Many resources are now generating, processing, storing, or providing kidney-related molecular, pathological, and clinical data. Reference ontologies offer an opportunity to support knowledge and data organization and integration. The Kidney Precision Medicine Project (KPMP) team contributed to the representation and addition of 329 kidney phenotype terms to the Human Phenotype Ontology (HPO) and identified many subcategories of acute kidney injury (AKI) or chronic kidney disease (CKD). The Kidney Tissue Atlas Ontology (KTAO) imports and integrates kidney-related terms from existing ontologies (e.g., HPO, CL, and Uberon) and represents 259 kidney-related biomarkers. We also developed a precision medicine metadata ontology (PMMO) to integrate 50 variables from KPMP and CellxGene resources and applied PMMO for integrative analysis. The gene expression profiles of kidney gene biomarkers were specifically analyzed in healthy controls or AKI/CKD disease states. This work demonstrates how ontology-based approaches support multi-domain data and knowledge organization and integration to advance precision medicine.

bioinformatics↗

bioRxiv2024

HSP60 chaperone deficiency disrupts the mitochondrial matrix proteome and dysregulates cholesterol synthesis

Mitochondrial proteostasis is critical for cellular function and survival. HSP60 is a molecular chaperone that interacts with more than 260 mitochondrial matrix proteins to assist in their folding and few genetic variants of HSP60 are compatible with life. The few reported human patients with HSP60 variants show phenotypes of neurodevelopmental delay associated with brain hypomyelination. It is currently unknown how deficiency of the HSP60 links to hypomyelination. Here, we studied the onset and progression of HSP60 deficiency in: (1) a HSP60 mutation-inducible cell system, (2) skin fibroblasts from patients with disease-associated HSP60 variants, and (3) zebrafish HSP60 knockout larvae. Collectively, we show how HSP60 deficiency leads to pervasive dysfunctions: (1) downregulated mitochondrial matrix proteome, (2) transcriptional activation of cytosolic stress responses, (3) and lipid accumulation with dysregulated cholesterol biosynthesis. In zebrafish larvae HSP60 deficiency induced early developmental abnormalities. Our comprehensive data identifies HSP60 as a master regulator of mitochondrial proteostasis and suggests a pivotal effect of HSP60 dysfunction on myelination through dysregulation of cholesterol biosynthesis. HighlightsO_LIHSP60 deficiency disrupts mitochondrial matrix proteins activating stress responses C_LIO_LIDisrupted matrix proteome impairs catabolism causing acetyl-CoA shortage C_LIO_LIHSP60 deficiency dysregulates cytosolic cholesterol synthesis C_LIO_LIHsp60 deficiency causes developmental abnormalities in zebrafish larvae C_LIO_LIDysregulated cholesterol synthesis links HSP60 deficiency to hypomyelination C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=59 SRC="FIGDIR/small/578131v1_ufig1.gif" ALT="Figure 1"> View larger version (11K): org.highwire.dtl.DTLVardef@891066org.highwire.dtl.DTLVardef@93ae5borg.highwire.dtl.DTLVardef@19bded0org.highwire.dtl.DTLVardef@414428_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2023

mTOR signaling controls the formation of smooth muscle cell-derived intimal fibroblasts during vasculitis.

The excessive accumulation of fibroblasts within the intimal layer of inflamed vessels is a feared complication of vasculitis, which can lead to arterial stenosis and ischemia. In this study, we have investigated how such intimal fibroblasts develop during Kawasaki Disease (KD), a paediatric vasculitis typically involving the coronary arteries. By performing lineage tracing studies in a murine model of KD, we reveal that vasculitis-induced intimal fibroblasts develop independently of both adventitial fibroblasts and endothelial cells, and instead derive from smooth muscle cells (SMCs). Notably, the emergence of SMC-derived intimal fibroblasts - in both mice and in patients with KD, Takayasus arteritis and Giant Cell arteritis - coincided with their activation of the mechanistic target of rapamycin (mTOR) signalling pathway. Moreover, the genetic deletion of mTOR signalling in SMCs abrogated the emergence of intimal fibroblasts, demonstrating that mTOR is an intrinsic and essential regulator of vasculitis-induced, SMC-derived intimal fibroblasts. Collectively these findings provide molecular insight into the pathogenesis of arterial stenosis and identify mTOR as a therapeutic target to prevent adverse vascular remodelling in vasculitis.

cell biology↗

bioRxiv2023

One-Hour Phosphoproteome Analysis with the Orbitrap Astral Mass Spectrometer

Owing to its roles in cellular signal transduction, protein phosphorylation plays critical roles in myriad cell processes. That said, detecting and quantifying protein phosphorylation has remained a challenge. We describe the use of a novel mass spectrometer (Orbitrap Astral) coupled with data-independent acquisition (DIA) to achieve rapid and deep analysis of human and mouse phosphoproteomes. With this method we map approximately 30,000 unique human phosphorylation sites within a half-hour of data collection. The technology was benchmarked to other state-of-the-art MS platforms using both synthetic peptide standards and with EGF-stimulated HeLa cells. We applied this approach to generate a phosphoproteome multi-tissue atlas of the mouse. Altogether, we detected 81,120 unique phosphorylation sites within 12 hours of measurement. With this unique dataset, we examine the sequence, structural, and kinase specificity context of protein phosphorylation. Finally, we highlight the discovery potential of this resource with multiple examples of novel phosphorylation events relevant to mitochondrial and brain biology.

systems biology↗