Biology subjects

Keller, S.

Publications and source records attributed to Keller, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Chemically induced USP21-KLHL12 association recruits USP21 to COPII assembly

Ubiquitin-specific protease 21 (USP21) is a deubiquitinase implicated in various biological processes and disease states, including cancer. To better understand USP21 protein interaction networks, we used biotin identification proximity labeling, and uncovered USP21 interactions with the cytoskeleton, endosomal, and transcriptional regulators. Notably, a subset of these interactions was modulated by the USP21 selective inhibitor BAY-805. In cells, BAY-805 promoted proximity between USP21 and Kelch like protein KLHL12, a component of the Cullin3 ligase complex. Structural modeling using AlphaFold3, together with hydrogen deuterium exchange mass spectrometry, identified an orthosteric BAY-805 binding site within USP21. Binding of BAY-805 functionally competes with ubiquitin at USP21, thereby enabling USP21 interaction with KLHL12. This BAY-805 induced proximity elicited relocalization of USP21 to KLHL12 containing coat protein complex II (COPII) vesicles. Induced USP21-KLHL12 association led to enlargement of COPII vesicles and impaired transport of large cargoes such as collagen. Overall, our findings reveal a novel chemically induced proximity mechanism in which the monovalent ligand BAY-805 outcompetes ubiquitin from USP21, thereby recruiting USP21 to KLHL12 associated protein complexes.

cell biology↗

bioRxiv2026

Membrane Thickness Strain from Protein Inclusions: A Multiscale Simulation and X-Ray Scattering Study of Proteoliposomes

Integral membrane proteins remodel the surrounding lipid bilayer, but quantifying the resulting deformations and linking them to protein density in the membrane has remained challenging. Here, we introduce an integrative methodology that combines all-atom molecular dynamics (MD) simulations with multiscale small-angle X-ray scattering (SAXS) analysis to connect membrane strain to the protein/lipid ratio in proteoliposomes. Using outer membrane phospholipase A (OmpLA) reconstituted into lipid bilayers with both increased and decreased hydrophobic thickness, we systematically probe the effects of positive and negative hydrophobic mismatch. MD simulations demonstrate that OmpLA causes anisotropic, oscillatory thickness deformations extending up to eight times the radius of the first lipid shell surrounding the protein, yet the net change in average membrane thickness remains below 1%. Through our multiscale SAXS analysis, we quantitatively extract structural parameters, ranging from proteoliposome size to internal membrane architecture, using constrained Bayesian inference, with priors derived from MD findings. Specifically, we determine the protein/lipid molar ratio and average membrane strain, revealing excellent agreement between experiment and simulation. In thinner bilayers, substantial protein loss limits the analysis, highlighting the role of bilayer stability in sample preparation. Moreover, the predominance of OmpLA monomers in the thicker membranes is consistent with weak, membrane-mediated repulsive interactions between protein inclusions. Collectively, this integrative approach establishes a framework for quantifying protein-lipid interactions across molecular and mesoscale dimensions.

biophysics↗

bioRxiv2026

An intestinal cell atlas and organoid model for the threespine stickleback

The intestine plays a crucial role in physiology, nutrition, and immune function, but intestinal anatomy and cell types have yet to be fully characterized in many fish species, the most diverse group of vertebrates. To address this gap, we characterized the structure and composition of the intestine of threespine stickleback (Gasterosteus aculeatus), an emerging model teleost in biological research. Using histology, myeloperoxidase staining, single-cell RNA sequencing, and RNA in situ hybridization, we defined major intestinal epithelial, immune, stromal, and stem/progenitor populations. Goblet cells were abundant in proximal and hindgut, while myeloperoxidase-positive granulocytes were evenly distributed throughout the intestine. To facilitate future experimental studies of stickleback intestinal function, we also developed the first intestinal organoid culture from stickleback and show that these cultures recapitulate epithelial architecture and retain expression of canonical intestinal epithelial markers. This organoid platform enables future functional studies of mucosal immunity, host-microbe interactions, and intestinal physiology in stickleback and related teleosts. Together, our integrated approach provides a comprehensive cell atlas and a novel experimental model for studying digestive and immune functions in threespine stickleback.

Cell Biology↗

bioRxiv2026

Genetic variation underlying plasticity in physiological traits mediates response to climate in a Populus hybrid zone

Phenotypic plasticity can buffer the potential fitness consequences of environmental change, yet limited understanding of its genetic basis constrains its application to predicting population response to future climates. Hybrid zones provide powerful systems to study the genetic basis of plasticity because admixture can create novel allele combinations that generate new reaction norms for selection to act upon. Here, we combine two clonally replicated common gardens of Populus trichocarpa x P. balsamifera genotypes with whole-genome resequencing to identify genetic variation underlying plasticity for physiological traits. Admixed genotypes exhibited broader, and in some cases novel, reaction norms relative to parental genotypes, particularly for key stomatal traits. Admixture mapping of genotype-specific reaction norms identified ten candidate genes on chromosome 15, including TWIST, associated with plasticity in adaxial stomatal occurrence and density. Using random forest models, we projected allele-specific responses to climate warming within the hybrid zone to link genetic variation in plasticity with predicted warming. Random forest models forecast that future climates would favor P. trichocarpa alleles at TWIST, while P. balsamifera alleles would be maintained in heterozygous genotypes. These results suggest that hybridization can expand reaction norms and maintain genetic variation that may facilitate rapid phenotypic response needed to adapt to climate change.

evolutionary biology↗

bioRxiv2026

Unveiling the taxonomic diversity and unprecedented biosynthetic treasure of the phylum Myxococcota

Natural products remain vital sources of therapeutics, particularly anti-infectives, and members of the phylum Myxococcota constitute an especially rich reservoir for their discovery. Based on decades of microbiological efforts, we present 154 new Myxococcota genomes and propose a revised taxonomy expanding the number of described families from 11 to 28 and genera from 32 to 90. Comparison with an equivalent set from the prime source Actinomycetota shows that Myxococcota possess a comparable biosynthetic diversity, underscoring their promise for large-scale isolation and sequencing efforts. The vast untapped potential reflected in 2,387 uncharacterized gene cluster families is highlighted by genome mining efforts, yielding four validated compounds exhibiting novel chemistry, including myxolutamids and myxopentacins. We show that many Myxococcota-derived natural products, such as myxolutamid A and two new sorangicin derivatives, are conserved within taxonomic lineages. New described families thus bear high biosynthetic potential underpinning the importance of precise taxonomic classification guiding targeted drug discovery.

microbiology↗

bioRxiv2025

StructGuy: Data leakage free prediction of functional effects of genetic variants.

The extent to which variations in protein-coding genes affect protein function has drawn the biological machine learning communitys attention to computationally model variant effect prediction tools. Multiplexed assays of variant effects (MAVE) experiments serve as a rich data source, but cannot deliver enough data for training truly large neural-net models. Therefore, zero-shot methods, for example protein language models, have increasingly gained popularity. For these methods, MAVE results serve primarily for evaluation purposes, as exemplified by the ProteinGym benchmark. In this study, we argue that the rapidly increasing amounts of MAVE data can be used to train efficient supervised methods, presenting our new tool StructGuy, based on gradient boosting trees methodology. In contrast to other supervised methods in the field, StructGuy, thanks to its dedicated training dataset and data leakage-free training process, can predict variant effects for proteins not seen during training. To evaluate this generalization ability, we constructed a dedicated benchmark and compared StructGuy with zero-shot methods from the ProteinGym leaderboard achieving a competitive performance. Further, we demonstrate that thanks to its architecture and careful feature engineering, we are able to provide fully interpretable predictions and direct explanations of the influence of mutations on protein three-dimensional structure, which favourably differs StructGuy from zero-shot tools.

bioinformatics↗

bioRxiv2025

Evolutionary dynamics of a lethal recessive allele in reintroduced fragmented lynx populations

Conservation programs worldwide use reintroductions and translocations as management tools to support endangered species. Such efforts often face genetic challenges, as small populations and restricted gene flow can lead to inbreeding and loss of genetic diversity. Inbreeding depression is a well-known risk in small, isolated populations, where reduced genetic diversity can compromise fitness and threaten population persistence. While correlations between inbreeding and reduced fitness are well documented, so far, no causal variant for a fitness-relevant phenotype has been described in a wild population. Here, we combine long-term health and population data with genomic analyses to identify a recessive lethal allele responsible for extensive organ mineralization in young Eurasian lynx (Lynx lynx) from a reintroduced population in Switzerland. The causal variant is located in FGF23 and predicted to impair secretion of fibroblast growth factor 23, an important regulator of phosphate homeostasis. The deleterious allele was likely introduced from one of the lynx source populations and subsequently increased in frequency through founder effects and inbreeding. All three Swiss lynx populations show high inbreeding levels (FROH > 0.4), reflecting substantial loss of genetic diversity. Our findings uniquely demonstrate how demographic history, founder effects, genetic drift, and inbreeding interact to shape the fate of a recessive deleterious allele, highlighting the importance of multiscale monitoring in wildlife conservation.

evolutionary biology↗

bioRxiv2025

Allosteric Gain-of-Function and Off-Mechanism Effects of ATP-Competitive Kinase Inhibitors Revealed by Multimodal Proteomics

ATP competitive kinase inhibitors represent one of the largest classes of targeted anti-cancer drugs. While their primary mechanism is to block catalytic activity, they can also trigger paradoxical phenotypic effects that cannot be explained by catalytic inhibition alone. These observations point to a hidden layer of drug action that modulates non-catalytic kinase functions via changes in kinase conformation and protein-protein interactions (PPIs). Here we developed a multimodal proteomics approach combining limited proteolysis coupled mass spectrometry on affinity purified samples (AP-LiP-MS), AP-MS, and proximity labeling-MS to map inhibitor-induced conformation and PPI changes. We show that inhibitor binding causes structural rearrangements in the AIDs of all tested kinases, consistent with a transition to an open, active-like kinase conformation. These structural shifts drive distinct kinase-protein interaction changes that control non-catalytic functions: sequestration of AMPK by inhibited CAMKK2 blocks phosphorylation by other kinases, CHEK1 inhibition causes dissociation from the mitochondrial protein CLPB and leads to mitochondrial fragmentation, and structural changes in inhibited PRKCA trigger rapid relocalization to cell junctions. Thus we identify the ATP-binding site as a major organizing center of kinase conformation and interaction. Our work suggests that these on-target, off-mechanism effects are highly prevalent and provides the analytical framework to systematically characterize a frequently overlooked phenomenon highly relevant for understanding drug side effects to guide the development of novel therapeutics.

molecular biology↗

bioRxiv2025

3D bone printing via primed differentiation of stem cells with ultrasound (3DBonUS)

Bone disorders and skeletal defects represent a significant clinical challenge. Tissue engineering and regenerative medicine (TERM) strategies using 3D bioprinting have emerged as promising alternatives, but still limited by the inability of directing stem cell differentiation in a controlled and reproducible manner. Advancing beyond this, we are proposing 3D bone printing via primed differentiation of stem cells with ultrasound (referred to here as 3DBonUS). This approach synergistically integrates low-intensity pulsed ultrasound (LIPUS) with a microfluidic-assisted 3D bioprinting platform enabling the biophysical stimulation of human bone marrow stromal cells (HBMSCs) during extrusion, promoting osteogenic differentiation without the need for post-fabrication treatments. Moreover, the incorporation of microbubbles enhanced the effects of LIPUS by amplifying mechanical signals at the cellular level. 3DBonUS was found to significantly upregulate key osteogenic markers (RUNX-2, ALP, COL1A1, BMP-2, OCN, OPN) as confirmed by immunofluorescence and RT-qPCR analysis. Furthermore, the LIPUS-treated constructs showed a significant increase in alkaline phosphatase activity and calcium deposition, indicating enhanced mineralisation. The 3DBonUS strategy represents a new modality in skeletal biofabrication, harnessing targeted minimally-invasive mechanical stimulation, with potential for manufacturing scalability and clinical application. Future studies will aim to validate 3DBonUS in vivo to assess the ultimate regenerative potential with enhanced osteogenic properties.

bioengineering↗

bioRxiv2025

Structure-function relationship of alpha-synuclein fibrillar polymorphs derived from distinct synucleinopathies

The aggregation of the protein alpha-synuclein (Syn) is a common feature of multiple neurodegenerative diseases collectively called synucleinopathies, for which the pathobiology is not well understood. The different phenotypic characteristics of the synucleinopathies Parkinsons disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA) have been proposed to originate from the distinct structures adopted by Syn in its amyloid forms. Here, using covalent labeling and limited proteolysis coupled to mass spectrometry (LiP-MS) in vitro and in situ within neuronal cells and directly in native patient brain homogenates, we show that pathogenic Syn from distinct synucleinopathies (PD, DLB and MSA) are structurally different. Further, we found that fibrillar structural differences are associated with different fibril interactomes and neuronal responses. We discovered disease-specific ubiquitination patterns and turnover profiles for pathogenic Syn species, detected molecular pathways responding specifically to the uptake of different Syn fibrillar polymorphs, and identified a subset of the involved proteins as candidate direct interactors of Syn. In particular, components of the Ubiquitin-proteasomal System (UPS), including E3 ubiquitin ligases, chaperones, and Deubiquitinating proteins, showed disease/polymorph-specific interaction patterns, possibly accounting for different resistance of patient-derived Syn fibrils to degradation. Genetic modulation with CRISPR-based tools showed that members of the UPS degradation pathway (three E3 ligases: UBE3A, TRIM25, HUWE1 and the AAA+ ATPase VCP) reduced Syn inclusions, in a strain-specific manner. LiP-MS also identified sets of proteins with altered protease susceptibility in postmortem brain homogenates of PD, DLB, and MSA patients. These sets were largely disease-specific and included proteins altered in cells treated with fibrils derived from patients with the matching disease. Our findings provide insight into cellular processes involved in the accumulation and turnover of Syn pathogenic aggregates in PD, DLB and MSA in a disease specific manner and constitutes a resource of potential novel drug targets in these synucleinopathies.

systems biology↗

bioRxiv2025

Water stable isotopes reveal the ecohydrological importance of stemflow for mature and juvenile European beech

O_LIStemflow of forest trees can contribute a significant fraction of water to forests water fluxes; however, it is still unclear if and to what extent trees use stemflow for water supply and how much stemflow is lost by percolating below the root zone. C_LIO_LIWe applied deuterium-enriched stemflow equivalent to a throughfall depth of 23 mm to adult Fagus sylvatica trees to trace stemflow through the soil and trees. We continuously measured in-situ water stable isotope compositions in soil and xylem water and destructively sampled xylem water in the crowns of adult labelled (n=18), unlabelled F. sylvatica (n=15) and unlabeled Picea abies (n=9), complemented by destructive xylem water sampling of neighboring juvenile F. sylvatica (n=45). C_LIO_LIStemflow water supported 3.9 - 14.0% of daily sap flux of adult labelled F. sylvatica trees. In the soil, deuterium-enriched stemflow was detectable at a max. of 0 - 0.40 m to the labelled tree. However, unlabeled juvenile trees within a distance of [~]2 m showed label water uptake, indicating rooting into soil compartments affected by stemflow label. C_LIO_LIWe demonstrate the importance of stemflow as a water source for both adult and neighboring juvenile F. sylvatica, strongly profiting from stemflow infiltration. C_LI

plant biology↗

bioRxiv2025

Brain structure-function coupling - relationship with language lateralisation

Language is one of the most extensively studied lateralised cognitive functions in the human brain, predominantly relying on the left hemisphere in most individuals. However, the mechanisms by which a stable white matter architecture underpins individual language functions remain unclear. Previous studies have employed structural connectivity (SC) and functional connectivity (FC) coupling for individual fingerprinting and task decoding, suggesting that variability in brain entropy may serve as a distinguishing characteristic for language lateralisation. We examined a large cohort of healthy adults (n = 285) to investigate SC-FC coupling and identify markers distinguishing different language laterality groups. Functional connectivity was measured using resting-state fMRI (rsfMRI) time-series data, while structural connectivity was determined via probabilistic fibre tractography. SC-FC coupling was investigated using the SENSAAS language atlas and defined as the Pearson correlation between non-zero elements of regional structural and functional connectivity profiles. Group differences were assessed using the PALM toolbox in FSL. Our findings revealed that increased SC-FC coupling in the left precentral sulcus was associated with typical language lateralisation, while increased coupling in the right middle temporal gyrus and left anterior insula was observed in individuals with atypical language lateralisation (pFDR < 0.05). Non-lateralised individuals exhibited increased coupling in the left anterior insula compared to lateralised (pFDR<0.05). SC-FC coupling offers a promising framework to uncover functional and anatomical differences among individuals with varying language lateralisation. This regional specificity indicates that typical, atypical, and non-lateralised profiles rely on different structural-functional alignments, likely reflecting the recruitment of alternative pathways for language processing.

neuroscience↗

bioRxiv2025

Atovaquone-venturicidin A combination is synergistic against Plasmodium falciparum

We report the synergistic interaction between the complex III inhibitor atovaquone and an ATP synthase inhibitor, venturicidin A, against Plasmodium falciparum. Our results suggest the simultaneous inhibition of the parasites primary and alternative pathways for generating the mitochondrial membrane potential to be responsible for this phenomenon, whereby the alternative pathway relies on ATP synthase running in reverse mode. We hypothesize that the synergistic interaction between atovaquone and proguanil could follow a similar mechanism.

microbiology↗

bioRxiv2025

Structural Plasticity of the Membrane-Bound Protein Degradation Assembly Supports Bacterial Adaptation to Stress

Protein degradation by AAA+ proteases is essential for bacterial adaptation to environmental stress. The membrane-bound AAA+ protease FtsH forms a large inner-membrane complex with the SPFH (Stomatin, Prohibitin, Flotillin, HflK/C) family transmembrane proteins HflK and HflC, playing a key role in bacterial recovery from aminoglycoside antibiotic stress. Recent structural studies have revealed both open, asymmetric and closed, symmetric conformations of the HflK/C assembly under different sample-preparation conditions, suggesting two distinct models for how this complex modulates FtsH proteolysis. To determine which conformation reflects the biologically active state, we engineered a disulfide-crosslinked HflK/C variant to stabilize the closed conformation and resolved its structure using high-resolution cryo-EM. Phenotypic assays showed that cells expressing either this stabilized, closed HflK/C variant or an HflK/C mutant that disrupts interactions with FtsH exhibit significantly impaired growth under aminoglycoside stress. Surprisingly, the cryo-EM structure of the FtsH*HflK/C complex from cells challenged with the aminoglycoside antibiotic tobramycin revealed a novel HflK/C arrangement, characterized by two openings on opposite sides that may facilitate substrate access to FtsH during proteotoxic stress. Together, our results suggest that both the dynamic open conformation of HflK/C and its specific interactions with FtsH are critical for adaptation to aminoglycoside-induced stress. Given the conserved structural and functional features of SPFH family members, our findings may offer a broader framework for understanding how this protein family operates under both basal and stress conditions.

biophysics↗

bioRxiv2025

Climate and hybridization shape stomatal trait evolution in Populus

O_LIStomata play a critical role in regulating plant responses to climate. Where sister species differ in stomatal traits, interspecific gene flow can influence the evolutionary trajectory of trait variation, with consequences to adaptation. C_LIO_LILeveraging six latitudinally-distributed transects spanning the natural hybrid zone between Populus trichocarpa-P. balsamifera, we used whole genome resequencing and replicate common garden experiments to test the role that interspecific gene flow and selection play to stomatal trait evolution. C_LIO_LIWhile species-specific differences in the distribution of stomata persist between P. balsamifera and P. trichocarpa, hybrids on average resembled P. trichocarpa. Admixture mapping identified several candidate genes associated with stomatal trait variation in hybrids including TWIST, a homolog of SPEECHLESS in Arabidopsis, that initiates stomatal development via asymmetric cell divisions. Geographic clines revealed candidate genes deviating from genome-wide average patterns of introgression, suggesting restricted gene flow and the maintenance of adaptive differences. Climate associations, particularly with precipitation, indicated selection shapes local ancestry at candidate genes across transects. C_LIO_LIThese results highlight the role of climate in shaping stomatal trait evolution in Populus and demonstrate how interspecific gene flow creates novel genetic combinations that may enhance adaptive potential in changing environments. C_LI

evolutionary biology↗

bioRxiv2025

White matter reorganization of motor and affective-motivational networks in pain-indifferent carriers of the R221W mutation

Congenital insensitivity to pain (CIP) due to the R221W mutation on the nerve growth factor gene results in reduced peripheral C-nociceptor density and behavioural indifference to painful stimuli. While functional neuroimaging has revealed altered cortical and sub-cortical pain processing in R221W carriers, structural white matter changes remain unexplored and may suggest an anatomical basis of symptoms. Heterozygous R221W carriers (n = 11) and age-, sex-, education-matched controls (n = 11) diffusion tensor imaging data were compared using fixel-based analysis, and complimentary edge and node analyses using graph theory, and network-based statistics. Whole-brain and region of interest (ROI) fixel-based analyses revealed significantly reduced fibre density and fibre-bundle cross-section in brainstem motor tracts of R221W carriers, encompassing the corticospinal pathways, corona radiata, external capsule, cerebellar peduncles, and pontine crossing (p < 0.05). Graph theory analysis of pain-processing ROIs demonstrated reduced local efficiency in right anterior cingulate cortex (ACC) and altered betweenness centrality in bilateral insula and left ACC of R221W carriers. Despite R221W carriers showing higher node degrees in the somatosensory cortex and ACC, these connections had reduced efficiency and integration with cortical network regions. Network-based statistics identified a possible compensatory subnetwork with stronger connectivity from right thalamus to left ACC and left insula in R221W carriers (p < 0.019). These findings suggest that congenitally reduced peripheral nociception could lead to abnormalities in the thalamocortical and motor efferent pathway, but not sensory afferent pathways. The combination of reduced brainstem motor tract integrity and altered cortical network efficiency, alongside potentially compensatory thalamo-cortical connectivity, could support a model of R221W CIP as motor under-reactivity rather than sensory insensitivity.

neuroscience↗

bioRxiv2025

Rapid adaptation and extinction across climates in synchronized outdoor evolution experiments of Arabidopsis thaliana

Climate change is threatening species with extinction, and rapid evolutionary adaptation may be their only option for population rescue over short ecological timescales. However, direct observations of rapid genetic adaptation and population dynamics across climates are rare across species. To fill this gap, we conducted a replicated, globally synchronized evolution experiment with the plant Arabidopsis thaliana for 5 years in over 30 outdoor experimental gardens with distinct climates across Europe, the Levant, and North America. We performed whole-genome sequencing on [~]70,000 surviving reproductive individuals and directly observed rapid and repeatable adaptation across climates. Allele frequency changes over time were parallel in experimental evolution replicates within the same climates, while they diverged across contrasting climates--with some allele frequency shifts best explained by strong selection between -46% to +60%. Screening the genome for signals of rapid climate adaptation identified a polygenic architecture with both known and novel adaptive genetic variants connected to important ecological phenotypes including environmental stress responses, CAM5 and HEAT SHOCK FACTORs, and germination and spring flowering timing, CYTOCHROME P450s and TSF. We found evolutionary adaptation trends were often predictable, but variable across environments. In warm climates, high evolutionary predictability was associated with population survival up to 5 years, while erratic trends were an early warning for population extinction. Together, these results show rapid climate adaptation may be possible, but understanding its limits across species will be key for biodiversity forecasting.

evolutionary biology↗

bioRxiv2025

The role of cytonuclear interactions to plant adaptation across a Populus hybrid zone

Co-adaptation of cytoplasmic and nuclear genomes are critical to physiological function for many species. Despite this understanding, hybridization can disrupt co-adaptation leading to a mismatch between maternally-inherited cytoplasmic genomes and biparentally inherited nuclear genomes. Few studies have examined the consequences of cytonuclear interactions to physiological function across environments. Here, we quantify the degree of co-introgression between chloroplast and nuclear-chloroplast (N-cp) genes across repeated hybrid zones and its consequences to physiological function across environments. We use whole-genome resequencing and common garden experiments with clonally replicated genotypes sampled across the natural hybrid zone between Populus trichocarpa and P. balsamifera. We use geographic clines to test for co-introgression of the chloroplast genome with N-cp and non-interacting nuclear genes. Co-introgression of chloroplast and N-cp genes was limited although contact zone-specific patterns suggest that local environments may influence co-introgression. Combining ancestry estimates with phenotypic data across common gardens revealed that mismatches between chloroplast and nuclear ancestry can influence physiological performance, but the strength and direction of these effects vary depending on the environment. Overall, this study highlights the importance of cytonuclear interactions to adaptation, and the role of environment in modifying the effect of those interactions.

evolutionary biology↗