Biology subjects

Williams, C.

Publications and source records attributed to Williams, C..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

SMARCA2 is an essential and potent cofactor for a specific subset of the glucocorticoid response in A549 cells

Glucocorticoids are a widely used, potent class of anti-inflammatory drugs that modulate the expression of hundreds of genes across the genome. Although the glucocorticoid response is primarily carried out by the glucocorticoid receptor (NR3C1, a.k.a. GR), there are many glucocorticoid receptor co-factors that are also essential to the downstream effects. To identify novel factors necessary for the glucocorticoid gene expression response, we used a genome-wide CRISPR screen in A549 lung adenocarcinoma cells. In that screen, we knocked out every gene in the human genome, and measured the effect of expression of the glucocorticoid-induced leucine zipper (GILZ), a classic glucocorticoid-response gene. We identified two chromatin remodeling proteins, SMARCA2 and BPTF, that are essential for GILZ expression. We then evaluated the genome-wide effects of SMARCA2 and BPTF on glucocorticoid-mediated gene expression. BPTF had a highly specific role in the glucocorticoid response, affecting the expression of only a handful of genes, and having virtually no effect on dexamethasone-induced changes in chromatin accessibility. However, SMARCA2 was necessary for 27% of dexamethasone-induced transcriptional changes (152 genes), and [~]7% of dexamethasone-induced changes in chromatin accessibility (586 regions of the genome). Genomic regions with SMARCA2-dependent changes in chromatin accessibility were characterized by high dexamethasone-induced regulatory activity in a massively parallel reporter assay, and dexamethasone-induced increases in transcription factor binding and chromatin states. Taken together, these data suggest that SMARCA2 is critical for chromatin remodeling at a specific set of genomic regions with high regulatory activity, which in turn drive changes in expression for many glucocorticoid-responsive genes.

genomics↗

bioRxiv2025

Multi-site Assessment of Methods for Cell Preservation Upstream of Single Cell RNA Sequencing

Single cell RNA sequencing (scRNA-seq) is a revolutionary technique to identify cell types and their molecular phenotype in heterogeneous biological specimens. ScRNA-seq typically requires fresh, high quality single cell suspensions that are processed immediately to preserve their molecular profiles. This presents a challenge for samples with long preparation times and prevents collection at remote sites lacking the required instrumentation for sample processing. Recently, several commercial assays have been released that enable sample preservation at the time of collection either via fixation or cryopreservation, allowing for sample processing to occur months after the initial collection. The Association of Biomolecular Research Facilities (ABRF) DNA Sequencing (DSRG) and Genomics Bioinformatics (GBiRG) Research Groups have undertaken a cross-platform, multi-site study to assess the performance and reproducibility of three platforms: a) 10x Genomics FLEX, b) Parse Bioscience Evercode WT v2 and c) Honeycomb Bio HIVE. Total leukocytes were isolated from a single healthy individual using the EasySep RBC depletion reagent. Cells were then characterized by collecting a 21-color flow cytometry dataset for reference and the remaining material was used for scRNA-seq procedures where different sites then processed either the fixed or cryopreserved cells for each method. We evaluated performance of each method across traditional scRNA-seq quality control metrics and analysis applications, including gene/transcript detection sensitivity, cell type discovery and annotation, and differential expression. We demonstrate that data from the methods tested can be effectively integrated and produce concordant results with regard to cell type annotation and relative abundance, though we observe platform-specific differences in the expression of a subset of genes. Preservation-based methods also show better retention of fragile granulocyte populations compared with fresh samples processed using the 10x 3 workflow. The improvements to preservation methods are changing the way research is conducted and our thorough investigation into the performance of each method provides a valuable resource to help scientists determine the most appropriate single cell preservation workflow given their sample collection logistics and laboratory infrastructure constraints.

genomics↗

bioRxiv2025

A live cell biosensor protocol for high-resolution screening of therapy-resistant cancer cells

The Genetically Encoded Death Indicator (GEDI) is a ratiometric, dual-fluorescence biosensor that enables real-time detection of cell death through calcium influx. Originally developed for use in neurodegeneration models, GEDI can be applied to cancer cells to quantify therapy-induced death at single-cell resolution. This protocol details how to generate GEDI-expressing cancer cell lines, empirically determine stress-induced GEDI thresholds using radiation or chemotherapeutic agents, and perform time-resolved imaging and image analysis to track cell fate. This workflow is optimized for high-throughput drug and radiation screening in heterogeneous populations and is especially useful for identifying chemo- and radio-resistant subclones. Key limitations include the need for empirical GEDI threshold calibration for each treatment condition and careful standardization of imaging parameters. The protocol outputs include GEDI ratio values, single-cell time-of-death annotations, and whole-cell morphological data in parallel, which can be linked to downstream applications such as FACS-based isolation of live or dying subpopulations, transcriptomic profiling of resistant clones, or in vivo validation using xenografts or organotypic slice culture. Associated contentio.protocol DOI: dx.doi.org/10.17504/protocols.io.eq2ly4d7qlx9/v1

cancer biology↗

bioRxiv2025

Gliomas phenocopy an inborn error of metabolism to drive neuronal activity and tumor growth

The metabolic hallmarks of high-grade glioma (HGG) are not fully understood. Human brain tissue metabolomics revealed that the creatine synthesis pathway intermediate guanidinoacetate (GAA) accumulated [~]100-fold in HGGs relative to controls, which was caused by imbalanced activities of enzymes in this pathway. Glioma cells secreted GAA rather than using it to produce creatine, implicating an unexpected function. GAA accumulates in GAMT deficiency, an inborn error of metabolism, and elevates neuronal excitability. Neuronal excitability is also increased in glioma and drives tumor growth through neuron-glioma interactions. We hypothesized that glioma-generated GAA excites surrounding neurons. Indeed, GAA induced neuronal hyperactivity by activating GABAA receptors and causing depolarizing GABA currents in glioma-associated neurons with dysregulated chloride homeostasis. Depleting tumoral GAA decreased electrochemical activity, neuron-glioma interactions, and tumor aggressiveness. Our findings unveil a new mechanism linking cancer metabolism with cancer neuroscience and leverage human genetics to nominate GAA synthesis as a target in gliomas.

cancer biology↗

bioRxiv2025

Design principles of a membrane-spanning ubiquitin ligase

Receptor-type E3 ubiquitin ligases are membrane-spanning assemblies that enable extracellular signals to directly control ubiquitylation in the cytoplasm. Despite playing widespread roles in tissue patterning and homeostasis, metabolism, and immunity, their structures and mechanisms remain poorly understood. Using cryo-electron microscopy, integrated with biophysical and functional studies, we visualized an E3 complex composed of two transmembrane proteins, MEGF8 and MOSMO, and the intracellular RING-family protein MGRN1. This MEGF8-MOSMO-MGRN1 (MMM) complex regulates left-right patterning of the body axis and the development of multiple organs, partly by attenuating signaling through the Hedgehog pathway. We find that the MMM complex functions like a fishing pole: a long, flexible helix attached to a membrane platform suspends an activated and precisely oriented RING domain--like a fishhook--to ubiquitylate the cytoplasmic surfaces of target receptors. Our structure explains how mutations in MEGF8 cause multi-organ birth defects in humans and defines a paradigm for receptor regulation by ubiquitylation.

biochemistry↗

bioRxiv2025

Lysosomal multi-omics reveals altered sphingolipid catabolism as driver of lysosomal dysfunction in the aging brain.

Recent data indicate that lipid composition has profound influence on the brain function and that changes in lipid homeostasis affect brain aging and predisposition to neurodegenerative diseases. Lipids dynamically reside in multiple intracellular locations and their organellar distribution is important for specific interactions and biological function. During brain aging lipid changes have been specifically noted in lysosomes, but the identity of the accumulated lipids, their interactions with other biomolecules such as proteins, and their functional relevance have not been characterized. We used mass spectrometry (MS) to assess longitudinal changes in the lipidome and proteome of lysosomes isolated from the mouse cortex, from the age of 3- to 24-months. Our statistical and machine learning analyses identified two factors demonstrating predictive power for age and differences in both lipids and proteins. Of these, factor 1 was the best predictor of sample age. Factor 1 lipids with the highest feature importance included multiple species of hexosylceramides (HexCer) and their sulfonated derivatives, sulfatides (SHexCer), all of which increased with age. Increased factor 1 proteins included myelin proteins, select sphingolipid catabolism enzymes and proteins associated with lysosomal storage diseases. Our analyses suggested that mechanisms underlying factor 1 encompass the combination of an age-dependent increase in lysosomal delivery of myelin components and alterations in lysosomal sphingolipid catabolism favoring degradation of sphingomyelin over HexCer. The overall age-related lysosomal changes resembled those observed in lysosomal storage diseases, particularly Gaucher disease, where accumulation of HexCer species is associated with lysosomal dysfunction. To corroborate factor 1 predictions, we employed a combination of biochemical, imaging and flow cytometry approaches, which confirmed alterations in sphingolipid catabolism and lysosomal accumulation of myelin components. These changes were associated with age-related alteration in lysosomal morphology, lysosomal dysfunction and inhibition of autophagy in both neurons and microglia. Our findings indicate that factors contributing to lysosomal aging resemble those observed in lysosomal storage diseases and underscore the significance of organelle-specific analyses for dissecting mechanisms contributing to brain aging.

neuroscience↗

bioRxiv2025

Le click c'est chic: a plug-and-play virus-like particle vaccination platform enabled by non-canonical amino acid incorporation and click chemistry in the tobacco BY-2 cell-free protein synthesis system

Non-canonical amino acids (ncaas) can provide recombinant proteins with novel exciting functionalities beyond the limits of nature, such as orthogonal reaction groups. Notably, ncaa introduction can be used in vaccinology to enhance the adaptability and immunogenicity of putative vaccine candidates. Cell-free protein synthesis (CFPS) represents the most promising methodology to introduce ncaa into recombinant proteins of interest. However, traditionally used prokaryotic CFPS systems show limitations to produce complex proteins requiring post-translational modifications, whilst eukaryotic CFPS systems have historically been difficult to scale and show low protein yields. In this work, we establish the site-specific introduction of ncaas into complex proteins with the high-yielding and scalable eukaryotic tobacco BY-2 CFPS system (BYL), commercialised as ALiCE(R). The tyrosine transferase from Escherichia coli (eTyrT) was tested for amber suppression-mediated ncaa incorporation in BYL. eTyrT showed high incorporation yields of up to 2mg/ml recombinant protein for the azido-tyrosine and alkyne-tyrosine ncaas, with linear scalability up to 10ml without any losses in protein yield. We applied ncaa incorporation in BYL to enable click chemistry bioconjugation of the receptor binding domain (RBD) of influenza hemagglutinin to pre-assembled hepatitis B core (HBc) virus-like particles (VLPs). BYL efficiently produced the alkyne-modified RBD and azido-modified HBc VLPs, and their conjugation via copper-catalysed azide-alkyne cycloaddition (CuAAC) led to structurally intact, RBD-coated particles. VLP-RBD conjugates could efficiently hemagglutinate chicken erythrocytes where the individual proteins could not, proving both the sialic-acid binding activity of the RBD and its multivalent presentation by the HBc VLP. Finally, when used to vaccinate mice the conjugated RBD-VLPs showed a greater protection against live influenza challenge than free RBD. This research thus enables ncaa introduction for recombinant proteins produced in BYL, constructing a novel plug-and-play vaccine platform and further expanding the capabilities of BYL to produce vaccine candidates and other proteins of interest.

molecular biology↗

bioRxiv2025

KeySDL: Sparse Dictionary Learning for Keystone Microbe Identification

Identification of microbes with large impacts on their microbial communities, known as keystone microbes, is a topic of long-standing interest in microbiome research. However, many approaches to identify keystone microbes are limited by the inherent nonlinearity and state-dependence of microbial dynamics. Machine learning approaches have been applied to address these shortcomings but often require more data than is available for a given microbial system. We propose a keystone identification approach called KeySDL which reduces the amount of data required by incorporating assumptions about the type of microbial dynamics present in the experimental system. The data are modeled as originating from a Generalized Lotka-Volterra (GLV) model, an architecture commonly used to simulate microbial systems. The parameters of this model are then estimated using Sparse Dictionary Learning (SDL) Compared to existing methods, this approach allows accurate prediction of keystone microbes from small numbers of samples and provides an output interpretable as reconstructed system dynamics. We also propose a self-consistency score to help evaluate whether the assumption of GLV dynamics is reasonable for a given dataset, either through the application of KeySDL or other analysis tools validated using GLV simulation.

bioinformatics↗

bioRxiv2025

Dynamic Culture Improves the Predictive Power of Bronchial and Alveolar Airway Models of SARS-CoV-2 Infection

Human in vitro lung models represent advanced tools for studying respiratory infections, particularly those caused by emerging respiratory pathogens. Despite scientific advances, vaccine and therapeutics pre-clinical development has yet to fully adopt human-relevant testing platforms due in part to a lack of validation. In this study, we characterised how static and dynamic flow culture conditions influence microphysiological systems (MPS) generated using primary bronchial and alveolar epithelial cells. We assessed epithelial structure, functional differentiation, and infection dynamics. This study represents the first direct comparison of how dynamic flow and endothelial co-culture influence viral tropism, replication kinetics, and host responses across anatomically distinct regions of the respiratory tract in vitro. Dynamic flow promoted formation of more physiologically relevant tissue architecture, pseudostratified bronchial epithelium and alveolar sac-like structures, with enhanced epithelial differentiation and retention of region-specific cell phenotypes at the transcriptomic level. Both static and dynamic flow models demonstrated responsiveness to inflammatory stimuli (poly(I:C), LPS), producing distinct, tissue-specific cytokine profiles and supporting infection with multiple SARS-CoV- 2 variants. Differences in infection efficiency, viral replication, and host gene expression were observed between variants, with dynamic flow models offering enhanced sensitivity and resolution. In alveolar tissues, dynamic flow increased infection efficiency and reduced variability, enabling more robust and consistent transcriptional responses. This facilitated the identification of interferon signalling pathways as key targets of the host response. Among the variants tested, Delta induced the most extensive tissue damage and strongest transcriptional response, whereas Omicron BA.5 exhibited greater infectivity in alveolar models compared to earlier variants. Our findings demonstrate that dynamic flow MPS more closely replicate human lung tissue architecture and cellular diversity, while also enhancing the predictive power and clinical relevance of airway models for ex vivo studies of SARS-CoV-2 infection. These improvements strengthen the reliability of data generated for the study of host-pathogen interaction studies and support the use of dynamic systems for evaluating novel anti-infectives, immunomodulators, and functional characterisation of immune sera generated by next-generation vaccines. Collectively, our results highlight the value of integrating dynamic in vitro models into preclinical pipelines for emerging respiratory pathogens.

microbiology↗

bioRxiv2025

Spatial analysis reveals a novel inflammatory tumor transition state which promotes a macrophage-driven induction of sarcomatoid renal cell carcinoma

Sarcomatoid renal cell carcinoma (sRCC) is an aggressive transdifferentiation of epithelioid clear cell RCC (ccRCC) tumors that shows heightened response to immunotherapy. The underlying biology leading to sarcomatoid transformation and mechanisms contributing to immunotherapy response are not well understood. Novel single cell spatial techniques were used in ccRCC and sRCC tumors from 40 patients to understand the spatial sRCC transformation and corresponding immune changes. A transcriptional transition state in epithelioid ccRCC cells along a continuum to mesenchymal sRCC was identified which expresses high levels of pro-inflammatory cytokines and an immune infiltrate. In vitro studies demonstrated that M2-like macrophages, recruited to the tumor by the transition state, induce full transition to the sarcomatoid state. A combination of increased PD-L1 expression and T-cells recruited by the transition state was observed, consistent with the increased immunotherapy response. This study enriches our understanding of the mechanisms leading to development and immune responsiveness of sRCC paving the way for novel approaches to diminish RCC progression.

cancer biology↗

bioRxiv2025

Contrasting roles for IKK regulated inflammatory signalling pathways for development and maintenance of type 1 and adaptive γδ T cells

The inhibitor of kappa B kinase complex (IKK) is a critical regulator of cell death and inflammatory signaling in multiple cell types. Phosphorylation of I{kappa}B proteins by IKK results in their degradation and consequent activation of NF-{kappa}B transcription factors. RIPK1, a critical cell death regulator, is also a direct target of IKK kinase activity, thereby repressing its cell death activity. In {beta} T cells, the RIPK1 kinase activity of IKK is critical for normal thymic development while mature {beta} T cells require IKK for both activation of NF-{kappa}B dependent survival programmes, and repression of RIPK1. {gamma}{delta} T cells play a unique and versatile role in host immunity with specific effector functions that enables them to act as early responders in immune defence. The role of IKK regulated pathways in their development and survival is not known. Here, we use mouse genetics to dissect the function of IKK and downstream pathways in the normal homeostasis of {gamma}{delta} T cells. We find that IKK expression is critical to establish a replete {gamma}{delta} T cell compartment, but that requires vary between different subsets. Type 1 {gamma}{delta} T cells require IKK dependent NF-{kappa}B activation for their generation, while IKK is redundant for development of adaptive {gamma}{delta} T cells. Instead, IKK dependent NF-{kappa}B activation is required for their longterm survival. We also find evidence that IKK repression of RIPK1 is required for survival of peripheral but not thymic {gamma}{delta} T cells. Ablation of CASPASE8 did not rescue {gamma}{delta} T cells in the absence of IKK but rather revealed a potent sensitivity of all {gamma}{delta} subsets to necroptosis, that was rescued by kinase dead RIPK1. Overall, we reveal critical requirements for IKK regulated inflammatory pathways by {gamma}{delta} T cells that contrast with those of {beta} T cells, and between different subsets, highlighting the complexity of the regulation of these pathways in the adaptive immune system.

immunology↗

bioRxiv2025

NMR Spectral Alignment Utilizing a CryoEM Motion Correction Algorithm

With recent advances in magic-angle spinning (MAS) solid-state NMR (SSNMR) resolution, precise spectral alignment has become a critical bottleneck in data processing workflows. While solution NMR employs deuterium lock systems, most SSNMR probes still lack this capability; though a lock corrects for magnet drift and instabilities, it is not alone sufficient to account for field gradients, sample temperature differences, and pulse sequence effects that can contribute to referencing errors among several data sets. These offsets become particularly problematic in the lengthy multidimensional experiments that provide the foundation for resonance assignment and structure determination procedures. Currently, researchers rely on manual alignment through visual peak inspection--a qualitative approach that often overemphasizes prominent, outlying peaks while overlooking subtle, global patterns. This subjective process becomes increasingly impractical for use cases with lower sensitivity, such as large proteins with thousands of peaks. To address these challenges, here we present Automated NMR Spectral Alignment (ANSA), a program that adapts cryo-electron microscopy motion correction principles to NMR spectroscopy. ANSA treats NMR spectra as images and applies cross-correlation functions to determine optimal alignment, improving cross-correlation scores from 0.33 to 1.00 in controlled tests and achieving 0.96 correlation in real-world applications with previously misaligned spectra. The algorithm successfully aligns spectra across varying experimental conditions, corrects shifts in long-duration experiments, and works with 2D and 3D datasets, with approaches that can be readily extended to additional dimensions. By eliminating human bias and providing objective, consistent spectral alignment, ANSA enhances scientific rigor, improves reproducibility between experiments, and enables automation of critical data processing steps. The software is freely available as an open-source tool, ready for integration into existing NMR workflows.

biophysics↗

bioRxiv2025

Ultra-High Resolution Solid-State NMR for High Molecular Weight Proteins on GHz-Class Spectrometers

NMR spectroscopy is a powerful technique with broad impact across the physical and life sciences, and ultra-high field, GHz-class NMR spectrometers offer exceptional overall performance including superior resolution and sensitivity. While the resolution is fundamentally limited by molecular tumbling for solution NMR, solid-state NMR (SSNMR) is constrained only by instrumentation, making it well-suited for studying large and complex systems. To fully leverage UHF magnets for magic-angle-spinning SSNMR, it is essential to eliminate linebroadening arising from magnetic field drift and couplings among the nuclear spins. We address these challenges using an external 2H lock to compensate the field drift and Long-Observation-Window Band-Selective Homonuclear Decoupling (LOW-BASHD) to suppress 13C homonuclear couplings. We thereby achieve better than 0.2 ppm resolution in proteins up to 144 kDa, enabling unique site resolution for over 500 amide backbone pairs in 2D experiments. This exceeds the resolution available from solution NMR for large biological molecules, greatly expanding the potential of GHz-class NMR for research in life sciences. TeaserUltra-high field NMR enables scientists to observe the finest details of large biomolecules, and this study overcomes key challenges to achieve a new benchmark for resolution in solid-state NMR of high molecular weight proteins.

biophysics↗

bioRxiv2025

Metabolic alterations in the absence of a detectable neuromuscular phenotype in novel genomically humanised SOD1A4V mice.

Amyotrophic lateral sclerosis (ALS) caused by mutation in superoxide dismutase 1 (SOD1) accounts for 15-30% of familial ALS and is typically autosomal dominant. How single base pair/amino acid changes in this small protein cause neurodegeneration is unknown. In North America, SOD1A4V is the most common familial ALS SOD1 mutation and results in an aggressive form of ALS. Here, we present a novel genomically humanised mouse model of SOD1A4V, in which the mouse Sod1 locus has been replaced by the human SOD1 gene, with intact genomic architecture of exons and introns, but bearing an A4V mutation. In agreement with previously reported human genomic knock-in mice, the phenotype is mild; however, transcriptomic and metabolomic profiling reveal significant dysregulation of glycolysis, the tricarboxylic acid (TCA) cycle, and lipid metabolism. These changes suggest an early bioenergetic imbalance that precedes neuromuscular impairment. Our findings support metabolic dysfunction as an early event in ALS pathogenesis. This freely available SOD1A4V model provides a valuable tool for studying ALS progression and identifying therapeutic targets for pre-symptomatic treatment. SUMMARY STATEMENTThis study describes the generation and analysis of novel genomically humanised SOD1A4V mice, revealing metabolic dysfunction through integrated multi-omic analyses, characterising a freely available potential pre-symptomatic ALS model for future research.

neuroscience↗

bioRxiv2025

The E3 ubiquitin ligase MGRN1 targets melanocortin receptors MC1R and MC4R via interactions with transmembrane adapters

E3 ubiquitin ligases play a crucial role in modulating receptor stability and signaling at the cell surface, yet the mechanisms governing their substrate specificity remain incompletely understood. Mahogunin Ring Finger 1 (MGRN1) is a membrane-tethered E3 ligase that fine-tunes signaling sensitivity by targeting surface receptors for ubiquitination and degradation. Unlike cytosolic E3 ligases, membrane-tethered E3s require transmembrane adapters to selectively recognize and regulate surface receptors, yet few such ligases have been studied in detail. While MGRN1 is known to regulate the receptor Smoothened (SMO) within the Hedgehog pathway through its interaction with the transmembrane adapter Multiple Epidermal Growth Factor-like 8 (MEGF8), the broader scope of its regulatory network has been speculative. Here, we identify Attractin (ATRN) and Attractin-like 1 (ATRNL1) as additional transmembrane adapters that recruit MGRN1 and regulate cell surface receptor turnover. Through co-immunoprecipitation, we show that ATRN and ATRNL1 likely interact with the RING domain of MGRN1. Functional assays reveal that MGRN1 requires these transmembrane adapters to ubiquitinate and degrade the melanocortin receptors MC1R and MC4R, in a process analogous to its regulation of SMO. Loss of MGRN1 leads to increased surface and ciliary localization of MC4R in fibroblasts and elevated MC1R levels in melanocytes, with the latter resulting in enhanced eumelanin production. These findings expand the repertoire of MGRN1-regulated receptors and provide new insight into a shared mechanism by which membrane-tethered E3 ligases utilize transmembrane adapters to dictate substrate receptor specificity. By elucidating how MGRN1 selectively engages with surface receptors, this work establishes a broader framework for understanding how this unique class of E3 ligases fine-tunes receptor homeostasis and signaling output.

cell biology↗

bioRxiv2025

Fibrillarin shapes oncogenic protein pools and ribosomal composition in triple-negative breast cancer

Fibrillarin (FBL), a core component of the C/D box snoRNP complex, catalyzes 2-O-methylation (Nm) of ribosomal RNA (rRNA), influencing ribosome heterogeneity and oncogene translation. In triple-negative breast cancer (TNBC), FBL dysregulation creates an aberrant Nm signature. This study explores role of FBL in TNBC progression via translation-driven mechanisms. FBL knockdown impaired tumorigenic traits, induced metabolic stress, and reduced the translation efficiency of oncogenes, including metastasis-associated protein 1 (MTA1), interleukin-1 receptor-associated kinase 1 (IRAK1), and thymosin beta 10 (TMSB10). RiboMethSeq analysis revealed differential rRNA Nm site sensitivity to FBL depletion. Additionally, FBL knockdown lowered RPS28 protein levels, suggesting its misincorporation into ribosomes. Notably, silencing RPS28 also suppressed oncogenic traits and downregulated MTA1, IRAK1, and TMSB10, highlighting its role in FBL-mediated translation. These findings uncover a complex interplay between FBL, rRNA Nm modifications, and RPS28 in shaping oncogenic protein pools and ribosomal composition in TNBC. Targeting this pathway could offer novel therapeutic strategies for this aggressive cancer subtype.

molecular biology↗

bioRxiv2025

Maternal undernutrition exacerbates microbiota-driven growth stunting through pre and postnatal effects

Linear growth stunting due to undernutrition affects 20% of children under the age of five with far-reaching consequences, including increased susceptibility to infection and altered cognitive development. Current nutritional interventions are largely ineffective in rescuing linear growth. A significant proportion of stunting originates in utero; however, the mechanisms by which maternal undernutrition is transmitted between generations remain poorly characterized. Here we employed a gnotobiotic murine model of intergenerational undernutrition in which offspring are exposed to distinct microbial communities both in utero and after birth. We found that maternal undernutrition exacerbated offspring growth deficits in a microbiota-dependent manner. Maternal undernutrition also altered the offspring microbiome, increasing abundance of Enterococcus species and Escherichia coli. Cross-fostering demonstrated critical roles for the microbiota in growth during both gestation and early life. These findings emphasize the need to target both mother and child in the design of nutritional therapies for undernutrition.

microbiology↗

bioRxiv2025

Functional, Immunogenetic, and Structural Convergence in Influenza Immunity between Humans and Macaques

Human B cell immunity to the influenza hemagglutinin (HA) stem region, a universal influenza vaccine target, is often stereotyped and immunogenetically restricted, posing challenges for study outside humans. Here, we show that macaques vaccinated with a HA stem immunogen elicit human-like public B cell lineages targeting two major conserved sites of vulnerability, the central stem and anchor epitopes. Central stem antibodies were predominantly derived from VH1-138, the macaque homolog of human VH1-69, a VH-gene preferentially used in human central stem broadly neutralizing antibodies (bnAbs). Similarly, macaques produced anchor bnAbs with the human-like NWP motif. Both bnAb lineages were functionally and structurally analogous to their human counterparts, with recognition mediated largely by germline-encoded motifs. Thus the macaque immunoglobulin repertoire supports human-like public bnAb responses to influenza HA. Moreover, this underscores the utility of homologous germline-encoded immunity, suggesting that immune repertoires of macaques and humans may have been similarly shaped during evolution. HIGHLIGHTS[bullet] Functional human-like public antibody lineages can be elicited to HA stem supersites in macaques. [bullet]Macaque central stem bnAbs are predominantly derived from VH1-138, a VH-gene homologous to human VH1-69. [bullet]The human-like CDR L3 NWP anchor epitope-targeting lineage can be elicited in macaques. [bullet]Central stem and anchor bnAbs from humans and macaques engage their respective epitopes with atomic level similarity.

immunology↗