Biology subjects

Mueller, M. C.

Publications and source records attributed to Mueller, M. C..

15 recordsLinked to original sources

bioRxiv2026

Translating Sex- and Disease-Informed Design Criteria from Human Lung into Engineered Biomaterials

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease characterized by pathological extracellular matrix (ECM) remodeling, and a pronounced sex disparity, with approximately 70% of cases occurring in males. Despite these differences, biomaterials used to model fibrosis are commonly designed to replicate disease state or stiffness alone and rarely account for biological sex. Here, we derived sex- and disease-informed biomaterial design criteria from female and male control and IPF human lungs and translated these criteria into engineered hybrid-hydrogels containing human lung decellularized ECM (dECM) and poly(ethylene glycol) alpha methacrylate (PEGMA). Matrisome-focused reanalysis of our previously published region-resolved human lung proteomic dataset revealed region- and sex-associated differences in fibrillar collagens, matricellular proteins, and structural proteoglycans. Complementary characterization of human lung tissue further identified sex- and disease-associated differences in fibrillar collagen architecture, proteoglycan and glycosaminoglycan (GAG) remodeling, and tissue mechanics. Male IPF lungs exhibited the highest levels of disease-associated stiffening and fibrillar collagen remodeling, while female IPF tissue showed comparatively modest mechanical changes despite biochemical remodeling. These measurements established tissue-informed design criteria spanning matrix composition, architecture, and mechanics. We next evaluated whether tissue-derived properties were reproduced or retained following decellularization, processing, and incorporation of donor-derived dECM into hybrid-hydrogels. Engineered hybrid-hydrogels closely recapitulated sex- and disease-associated mechanical relationships, while COL1A1-associated abundance patterns were more consistently preserved than fibrillar collagen organization. Matricellular proteins and GAG-rich matrix remained detectable, although condition-dependent biochemical fidelity varied. This work establishes a human tissue-informed framework for engineering sex- and disease-specific biomaterials, highlighting the importance of defining sex-associated ECM properties and validating individual features following material fabrication to advance human models of IPF and other diseases with sex-associated differences in susceptibility and progression.

bioengineering↗

bioRxiv2026

Dynamic hybrid-hydrogel lung models decouple matrix composition, stiffness, and fibroblast memory to define distinct drivers of fibrotic progression

Idiopathic pulmonary fibrosis is a devastating chronic lung disease characterized by progressive scarring of the lung, which leads to impaired gas exchange and ultimately death. While research has provided us with extensive understanding on end-stage disease, the factors that lead to forward-feedback loops of fibrotic progression are still not fully known. Cell intrinsic activation, pathological extracellular matrix (ECM) composition, and increased tissue stiffness are all hallmarks of advanced fibrosis, but the relative contribution of these factors to disease has been difficult to disentangle using classic in vivo models. In this study we created biomaterials-based 3D lung models that incorporate geometrically relevant co-culture of lung epithelial cells and fibroblasts with tunable stiffness, ECM-containing hybrid-hydrogels. Using this model system, we demonstrated that environmental stiffness has the strongest effect on overall fibroblast activation. RNAseq analysis revealed unique gene-level changes in both fibroblasts and epithelial cells due to both composition and stiffness, highlighting the importance of incorporating both factors into any in vivo disease models. Overall, these results reinforce the value of biomaterials-based models in understanding disease pathogenesis, and their potential for screening of treatment responses.

bioengineering↗

bioRxiv2026

Proteomic analysis reveals regional sex differences in healthy and fibrotic human lung

Biological sex has systemic effects on gene expression, cell behavior, and disease etiology. Despite these widespread effects, sex as a biological variable is understudied, particularly in chronic lung diseases. In idiopathic pulmonary fibrosis (IPF), 70% of patients are male, and male patients have overall worse survival post-diagnosis. While behavioral differences between sexes might account for some of the epidemiological differences, the contribution of underlying biology is not known. In this study, we performed regional proteomic analysis via laser-captured microdissection-coupled mass spectrometry and analyzed the data for sex-biased protein expression. We discovered that even in control lung, sex differences existed in both airway and alveolar regions. Sex differences became more pronounced in diseased regions, with sex-biased expression of diverse proteins including those involved in extracellular vesicle secretion, cellular metabolism, and extracellular matrix remodeling. These data suggest that baseline sex differences in lung proteome may contribute to sex-specific susceptibility, progression, and clinical outcomes in IPF, underscoring the need for future mechanistic and clinical studies to account for sex as a biological variable.

bioengineering↗

bioRxiv2026

Proteogenomics of Blumeria hordei supports RNA and protein coding innovative potential derived from transposable elements

Some filamentous plant-pathogenic fungi have comparably large genome sizes within the fungal kingdom due to the proliferation of transposable elements (TEs). Blumeria hordei (Bh), the causal agent of the powdery mildew disease on barley, is a filamentous obligate biotrophic fungus. Compared to other ascomycetes, it contains a low number of genes but a high genomic TE content of approximately 75%. Yet, a comprehensive understanding of the contribution of TEs to the RNA and protein landscape of Bh is lacking. Here, we use Bh as a model to study transcripts and proteins derived from genes and individual TEs. Therefore, we created two high-quality genome assemblies of the German Bh isolate TUM1 and the Australian Bh isolate AUS1. We applied deep proteomics with mass spectrometry, long-read and short-read sequencing on both DNA and RNA. Based on these multi-omic resources, we completed nearly gapless genome assemblies, new gene and TE annotations, and effector predictions. Using long-read RNA sequencing, we detected extensive co-transcription of TEs and genes as TE-gene chimeric transcripts. We identified previously unpredicted splice variants or genes, partially supported by proteomics. The intergenic and TE genomic space of Bh TUM1 gives rise to thousands of transcripts and several novel TE-derived proteins that lack from previous TE protein predictions. Together, this supports an existing potential for expression of novel transcripts and proteins from highly abundant TEs in the Bh genome.

plant biology↗

bioRxiv2026

The crop pathogen Blumeria hordei exhibits genome-wide pervasive selective and neutral sweepstakes reproduction signatures.

The fungal crop pathogen Blumeria hordei, causal agent of powdery mildew on barley, presents life-history and epidemiological characteristics, as well as and selective pressures due to modern agriculture leading to expected sweepstakes reproduction, that is highly skewed offspring distributions. Using genome-wide polymorphism data and population genomics inferences, we aim to 1) infer the past demographic history and the strength of sweepstakes reproduction in B. hordei, and 2) quantify the contributions of these selective and neutral processes in the genome. An new inference method based on Neural Posterior Estimation and diversity and linkage disequilibrium statistics was developed and tested on simulated and B. hordei genomic data. We confirm that B. hordei exhibits a moderate sweepstakes reproduction (-parameter of 1.6). We highlight that the Site Frequency Spectrum (SFS) appears sensitive to the joint occurrence of sweepstakes and recent demographic changes, which may caution on the reliability of the SFS to infer sweepstakes reproduction. We then scan the genome for selective sweeps, adjusting the significance thresholds of the methods for demographic history and sweepstakes reproduction, thereby yielding a counterintuitive result. When conditioning the significance threshold for sweep detection on simulations under sweepstakes and demography, a very large number of putatively selected regions is found (11.6% of the genome). We suggest that sweepstakes reproduction in B. hordei is due to 1) neutrality (clonal/sexual phases and Boom-and-Bust cycles) generating a genome-wide level of background noise in the coalescent genealogies, and 2) selective sweepstakes due to pervasive positive selection. Our findings have important implications for both population genomic methodology and our understanding of pathogen evolution.

evolutionary biology↗

bioRxiv2026

Engineering quantitative root disease resistance in barley by targeting conserved SCAR susceptibility genes without compromising seed yield or mycorrhizal symbiosis

Durable resistance to soil-borne pathogens remains elusive in cereals, partly because susceptibility (S) genes that facilitate root infection have not been identified in monocots. In the model legume Medicago truncatula, the SCAR/WAVE complex member MtAPI functions as a root S-gene for microbial invasion. Whether SCAR gene associated susceptibility function is conserved in monocots, and whether SCAR gene inactivation can enhance root resistance in cereals, remains unknown. Here, we identify and characterize three SCAR genes in barley: HvSCAR-A, HvSCAR-B, and HvSCAR-C. Cross-species complementation assays indicate that HvSCAR-B and HvSCAR-C are functionally similar to MtAPI. While hscar-b and hvscar-c single mutants exhibited no major growth defects, hvscar-a mutants showed strongly reduced seed production, and a hvscar-b/c double mutant displayed shorter root hairs. Notably, the hvscar-b/c double mutant exhibited increased resistance to the hemibiotrophic pathogen Phytophthora palmivora but greater colonization by the symbiotic arbuscular mycorrhizal fungus Funneliformis mosseae, underscoring a complex role in plant root - microbe interactions. Our findings reveal a conserved susceptibility function of SCAR genes in monocots and identify api monocot homologs as promising targets for engineering disease resistance in cereals. This study offers new insights into SCAR protein functional diversification and its potential for improving root health in crop plants.

plant biology↗

bioRxiv2025

Dual recognition of structurally unrelated mildew effectors underlies the broad-spectrum resistance of Pm3e in wheat

Broad-spectrum resistance genes are highly valuable for sustainable crop protection, yet the molecular basis of their activity is often unknown. The Pm3 allelic series in wheat encodes NLR receptors that recognize avirulence (AVR) effectors of wheat powdery mildew. Here, we show that near-identical Pm3 alleles vary greatly in resistance efficacy and broadness against a global mildew isolate collection and subsequently use this model system to study the mechanisms underlying broad-spectrum resistance. We demonstrate that two alleles, Pm3d and Pm3e, provide resistance against the majority of isolates worldwide, by each recognizing two AVR effectors from powdery mildew, thereby lowering the risk of resistance breakdown. While Pm3d recognizes two closely related RNase-like AVR effectors, Pm3e detects two structurally diverse AVRs, including an effector belonging to a large, uncharacterized protein family with a novel structural fold. Using chimeric Pm3 NLRs, we identify specificity-defining polymorphisms of Pm3d and Pm3e against their diverse effector targets. Lastly, we demonstrate that Pm3d and Pm3e activities can be combined in engineered Pm3 NLRs, thereby further extending their recognition spectrum. Our findings highlight the potential of Pm3 immune receptors for long-lasting wheat protection by demonstrating their versatility in recognizing structurally diverse effectors and their amenability to NLR engineering.

plant biology↗

bioRxiv2025

An HMA-like integrated domain in the wheat tandem kinase WTK4 recognises an RNase-like pathogen effector

Proteins with a tandem kinase structure have recently emerged as new players in race-specific resistance in cereal crops. However, the molecular understanding of these novel immune receptors resistance mechanisms is limited by the lack of knowledge about the pathogen effectors that they recognise. In this work, we identify AvrWTK4, the wheat powdery mildew RNase-like effector recognised by the wheat tandem kinase immune receptor WTK4, through a combination of bi-parental genetic mapping and mutagenesis. We demonstrate that mutations in the AvrWTK4 gene or a reduction of its expression lead to virulence on WTK4. Transfection of AvrWTK4 specifically induced cell death in WTK4-containing Aegilops tauschii protoplasts. The avirulent AvrWTK4 variant interacts more strongly than the virulent variant with the N-terminal heavy metal-associated (HMA)-like domain of WTK4. These findings further highlight that integrated domains in tandem kinase proteins serve as decoys for pathogen effectors, which could be leveraged to design novel recognition specificities.

molecular biology↗

bioRxiv2025

Virulence on Pm4 kinase-based resistance is determined by two divergent wheat powdery mildew effectors

The wheat resistance gene Pm4 encodes a kinase fusion protein and has gained particular attention as it confers race-specific resistance against two major wheat pathogens: powdery mildew and blast. Here, we describe the identification of AvrPm4, the mildew avirulence effector recognised by Pm4, using UV- mutagenesis, and its functional validation in wheat protoplasts. We show that AvrPm4 directly interacts with and is phosphorylated by Pm4. Using genetic association and QTL mapping, we furthermore demonstrate that evasion of Pm4 resistance by virulent mildew isolates relies on a second fungal component, SvrPm4, which suppresses AvrPm4-induced cell death. Surprisingly, SvrPm4 was previously described as AvrPm1a. We show that SvrPm4, but not its inactive variant svrPm4, is recognised by the NLR immune receptor Pm1a. These multiple roles of a single effector provide a new perspective on fungal (a)virulence proteins and their combinatorial interactions with different types of immune receptors.

genetics↗

bioRxiv2025

Female fibroblast activation is estrogen-mediated in sex-specific 3D-bioprinted pulmonary artery adventitia models

Pulmonary arterial hypertension (PAH) impacts male and female patients in different ways. Female patients exhibit a greater susceptibility to disease (4:1 female-to-male ratio) but live longer after diagnosis than male patients. This complex sexual dimorphism is known as the estrogen paradox. Prior studies suggest that estrogen signaling may be pathologic in the pulmonary vasculature and protective in the heart, yet the mechanisms underlying these sex-differences in PAH remain unclear. PAH is a form of a pulmonary vascular disease that results in scarring of the small blood vessels, leading to impaired blood flow and increased blood pressure. Over time, this increase in blood pressure causes damage to the heart. Many previous studies of PAH relied on male cells or cells of undisclosed origin for in vitro modeling. Here we present a dynamic, 3D-bioprinted model that incorporates cells and circulating sex hormones from female patients to specifically study how female patients respond to changes in microenvironmental stiffness and sex hormone signaling. Poly(ethylene glycol)-alpha methacrylate (PEGMA)-based hydrogels containing female human pulmonary artery adventitia fibroblasts (hPAAFs) from idiopathic PAH (IPAH) or control donors were 3D bioprinted to mimic pulmonary artery adventitia. These biomaterials were initially soft, like healthy blood vessels, and then stiffened using light to mimic vessel scarring in PAH. These 3D-bioprinted models showed that stiffening the microenvironment around female IPAH hPAAFs led to hPAAF activation. On both the protein and gene-expression levels, cellular activation markers significantly increased in stiffened samples and were highest in IPAH patient-derived cells. Treatment with a selective estrogen receptor modulator reduced expression hPAAF activation markers, demonstrating that hPAAF activation is a one pathologic response mediated by estrogen signaling in the vasculature, validating that drugs currently in clinical trials could be evaluated in sex-specific 3D-bioprinted pulmonary artery adventitia models.

bioengineering↗

bioRxiv2024

Genomic surveillance and molecular evolution of fungicide resistance in European populations of wheat powderymildew

Fungicides are used in agriculture to manage fungal infections and maintain crop yield and quality. In Europe, their application on cereals increased drastically starting from the mid 1970s, contributing to a significant improvement in yields. However, extensive usage has led to the rapid evolution of resistant pathogen populations within just a few years of fungicide deployment. Here we focus on wheat powdery mildew, a disease caused by the ascomycete fungus Blumeria graminis forma specialis tritici (Bgt). Previous research on Bgt documented the emergence of resistance to different fungicides and identified various resistance mechanisms. Yet, the frequency, distribution, and evolutionary dynamics of fungicide resistance in Bgt populations remain largely unexplored. In this study we leveraged extensive sampling and whole-genome sequencing of Bgt populations in Europe and the Mediterranean to investigate the population genetics and molecular epidemiology of fungicide resistance towards five major fungicide classes. We analyzed gene sequences and copy number variation of eight known fungicide target genes in 415 Bgt isolates sampled between 1980 and 2023. We observed that mutations conferring resistance to various fungicides increased in frequency over time, and had distinct geographic distributions, likely due to diverse deployment of fungicides across different regions. For demethylation inhibitor fungicides we identified multiple independent events of resistance emergence with distinct mutational profiles, and we tracked their rapid spread in the last decades. Overall, we revealed the evolutionary and epidemiological dynamics of fungicide resistance mutations in European Bgt populations. These results underscore the potential of genomic surveillance and population genetics to enhance our understanding of fungicide resistance.

pathology↗

bioRxiv2024

Avirulence depletion assay: combining R gene-mediated selection with bulk sequencing for rapid avirulence gene identification in wheat powdery mildew

Wheat production is threatened by multiple fungal pathogens, such as the wheat powdery mildew fungus (Blumeria graminis f. sp. tritici, Bgt). Wheat resistance breeding frequently relies on the use of resistance (R) genes that encode diverse immune receptors which detect specific avirulence (AVR) effectors and subsequently induce an immune response. While R gene cloning has accelerated recently, AVR identification in many pathogens including Bgt lags behind, preventing pathogen-informed deployment of resistance sources. Here we describe a new "avirulence depletion (AD) assay" for rapid identification of AVR genes in Bgt. This assay relies on the selection of a segregating, haploid F1 progeny population on a resistant host, followed by bulk sequencing, thereby allowing rapid avirulence candidate gene identification with high mapping resolution. In a proof-of- concept experiment we mapped the AVR component of the wheat immune receptor Pm3a to a 25kb genomic interval in Bgt harboring a single effector, the previously described AvrPm3a2/f2. Subsequently, we applied the AD assay to map the unknown AVR effector recognized by the Pm60 immune receptor. We show that AvrPm60 is encoded by three tandemly arrayed, nearly identical effector genes that trigger an immune response upon co- expression with Pm60 and its alleles Pm60a and Pm60b. We furthermore provide evidence that Pm60 outperforms Pm60a and Pm60b through more efficient recognition of AvrPm60 effectors, suggesting it should be prioritized for wheat breeding. Finally, we show that virulence towards Pm60 is caused by simultaneous deletion of all AvrPm60 gene paralogs and that isolates lacking AvrPm60 are especially prevalent in the US thereby limiting the potential of Pm60 in this region. The AD assay is a powerful new tool for rapid and inexpensive AVR identification in Bgt with the potential to contribute to pathogen-informed breeding decisions for the use of novel R genes and regionally tailored gene deployment.

plant biology↗

bioRxiv2023

Dynamic Microenvironmental Stiffness and Serum Composition Modulate Sex-Specific Pulmonary Artery Adventitial Fibroblast Activation

Respiratory diseases like pulmonary arterial hypertension (PAH) frequently exhibit sexual dimorphism. Female PAH patients are more susceptible to the disease but have increased survival rates. This phenomenon is known as the estrogen paradox, and the underlying mechanisms are not fully understood. During PAH progression in vivo, human pulmonary arterial adventitial fibroblasts (hPAAFs) differentiate into an activated phenotype. These cells produce excess, aberrant extracellular matrix proteins that stiffen the surrounding pulmonary arterial tissues. Here, we employed dynamic poly(ethylene glycol)-alpha methacrylate (PEGMA)-based biomaterials to study how the age and sex of human serum influenced hPAAF activation in response to microenvironmental stiffening in vitro. Results showed female and male cells responded differently to increases in microenvironmental stiffness and serum composition. Male hPAAFs were less activated than female cells on soft hydrogels and more responsive to increases in microenvironmental stiffness regardless of serum composition. Female hPAAF activation followed this pattern only when cultured in younger (age < 50) female serum or when older (age [≥] 50) female serum was supplemented with estradiol. Otherwise, female hPAAF activation was relatively high on both soft and stiffened hydrogels, with little difference in activation between the two conditions. Collectively, these results suggest that it may be possible to model the estrogen paradox observed in PAH in vitro and that it is critical for researchers to report cell sex and serum source when conducting in vitro experimentation.

bioengineering↗

bioRxiv2022

Doomed by popularity: The broad use of the Pm8 resistance gene in wheat resulted in hypermutation of the AvrPm8 gene in the powdery mildew pathogen

O_LIThe Pm8 resistance gene against the powdery mildew disease has been introgressed from rye into wheat as part of a large 1BL.1RS chromosomal translocation. Due to its high agronomic value, this translocation has seen continuous global use since the 1960s on large growth areas, even after Pm8 resistance was overcome. This allows studying the effect of long and widespread resistance gene use on a pathogen population. C_LIO_LIUsing genome wide association studies in a global population of wheat mildew isolates, we identified the avirulence effector AvrPm8 specifically recognized by Pm8. C_LIO_LIHaplovariant mining in the global population revealed 17 virulent haplotypes of the AvrPm8 gene that grouped into two categories. The first one from geographically diverse regions comprised two single amino acid polymorphisms at the same position in the AvrPm8 protein, which we confirmed to be crucial for recognition by Pm8. The second category consisted of numerous destructive mutations to the AvrPm8 open reading frame. C_LIO_LIMost individual gain-of-virulence mutations were found in geographically restricted regions, indicating they occurred recently as a consequence of the frequent Pm8 use. We conclude that both standing genetic variation as well as locally occurring new mutations contributed to the global breakdown of the Pm8 resistance gene introgression. C_LI

plant biology↗

bioRxiv2021

Standing genetic variation of the AvrPm17 avirulence gene in powdery mildew limits the effectiveness of an introgressed rye resistance gene in wheat

Introgressions of chromosomal segments from related species into wheat are important sources of resistance against fungal diseases. The durability and effectiveness of introgressed resistance genes upon agricultural deployment is highly variable - a phenomenon that remains poorly understood as the corresponding fungal avirulence genes are largely unknown. Until its breakdown, the Pm17 resistance gene introgressed from rye to wheat provided broad resistance against powdery mildew (Blumeria graminis). Here, we used QTL mapping to identify the corresponding wheat mildew avirulence effector AvrPm17. It is encoded by two paralogous genes that exhibit signatures of re-occurring gene conversion events and are members of a mildew sub-lineage specific effector cluster. Extensive haplovariant mining in wheat mildew and related sub-lineages identified several ancient virulent AvrPm17 variants that were present as standing genetic variation in wheat powdery mildew prior to the Pm17 introgression, thereby paving the way for the rapid breakdown of the Pm17 resistance. QTL mapping in mildew identified a second genetic component likely corresponding to an additional resistance gene present on the 1AL.1RS translocation carrying Pm17. This gene remained previously undetected due to suppressed recombination within the introgressed rye chromosomal segment. We conclude that the initial effectiveness of 1AL.1RS was based on simultaneous introgression of two genetically linked resistance genes. Our results demonstrate the relevance of pathogen-based genetic approaches to disentangle complex resistance loci in wheat. We propose that identification and monitoring of avirulence gene diversity in pathogen populations becomes an integral part of introgression breeding to ensure effective and durable resistance in wheat. Significance StatementDomesticated and wild wheat relatives provide an important source of new immune receptors for wheat resistance breeding against fungal pathogens. The durability of these resistance genes is variable and difficult to predict, yet it is crucial for effective resistance breeding. We identified a fungal effector protein recognised by an immune receptor introgressed from rye to wheat. We found that variants of the effector allowing the fungus to overcome the resistance are ancient. They were already present in the wheat powdery mildew gene pool before the introgression of the immune receptor and are therefore responsible for the rapid resistance breakdown. Our study demonstrates that the effort to identify new resistance genes should be accompanied by studies of avirulence genes on the pathogen side.

plant biology↗