Biology subjects

Keller, A.

Publications and source records attributed to Keller, A..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Multimodal brain cell atlas across the adult macaque lifespan

High-throughput single-cell omics of non-human primate tissues present a remarkable opportunity to study primate brain aging. Here, we introduce a transcriptomic and chromatin accessibility landscape of 1,985,317 cells from eight brain regions of 13 cynomolgus female monkeys spanning adult lifespan including exceptionally old individuals up to 29-years old. This dataset uncovers dynamic molecular changes in critical brain functions such as synaptic communication and axon myelination, exhibiting a high degree of cell type and brain region specificity. We identify the multicellular networks of the pons and medulla as a previously unrecognized hotspot for aging. Furthermore, comparative analyses with human neurodegeneration datasets highlight both shared and distinct mechanisms contributing to aging and disease. In addition, we uncover transcription factors implicated in monkey brain aging and pinpoint aging-regulated loci linked to longevity and neurodegeneration. This spatiotemporal atlas will advance our understanding of primate brain aging and its broader implications for health and disease.

neuroscience↗

bioRxiv2025

Improving allometric models to estimate the proboscis length of tropical bees

The proboscis length of bees is a key morphological trait shaping communities, pollination networks, and likely their responses to habitat loss. Despite its importance, it is rarely considered in ecological studies because of logistic limitations in obtaining accurate measurements across many different species. In two previous studies, the proboscis length of temperate bee species was estimated based on body size and bee family. However, bee taxa partially occurring in the tropics might deviate from this allometric relationship due to different functional constraints. Thus, we tested if equations developed for temperate bees can accurately predict the proboscis length in Meliponini, Euglossini (both Apidae), and Augochlorini (Halictidae), three ubiquitous and highly important tribes of tropical bees. We measured the intertegular distance (as a proxy of body size measurement) and the proboscis length of 892 specimens of 105 tropical species. We used these measurements to evaluate the previous model and found that its estimations lacked accuracy when applied to tropical bees, particularly to Meliponini and Euglossini. We developed new allometric equations estimating the proboscis length based on the intertegular distance, using (sub-) genera as an additional predictive variable to refine the estimations. We tested our approach by creating a test model for Meliponini, trained with only 80 % of the data, and evaluated this model using the remaining 20 %, resulting in a high accuracy of estimates. Our results shed additional light on the nature of the proboscis length-body size allometric relationship in tropical bees and provide a tool for future studies on the functional ecology of bees and their interactions with plants.

zoology↗

bioRxiv2025

Gestational Chlorpyrifos Exposure Imparts Lasting Alterations to the Rat Somatosensory Cortex

Chlorpyrifos is an organophosphorus pesticide used extensively in agricultural and residential settings for nearly 60 years. Gestational, sub-acute exposure to chlorpyrifos is linked to increased prevalence of neurodevelopmental disorders. Animal studies have modeled these neurobehavioral detriments, however, the functional alterations in the brain induced by this exposure remain largely unknown. To address this, we used a rat model of gestational chlorpyrifos exposure to interrogate the alterations in the developing somatosensory (barrel) cortex. Rat dams were exposed to chlorpyrifos (5 mg/kg) or vehicle on gestational days 18-21 via subcutaneous injection, with no overt acute toxicity. Acetylcholinesterase was modestly inhibited but returned to baseline levels by postnatal day 12. We performed whole-cell patch clamp recordings on postnatal days 12-20 in both male and female progeny of the treated dams. A spike timing dependent plasticity protocol revealed changes to the normal development of use-dependent plasticity, including interference in long-term synaptic depression. Recording inhibitory synaptic activity revealed an increase in the frequency of spontaneous postsynaptic currents and in paired pulse ratios, in conjunction with a significant decrease in miniature postsynaptic currents. These findings suggest a presynaptic mechanism of inhibited GABA release, with potential disinhibition of inhibitory neurons. Evaluation of barrel cortex development displayed disruptions to normal barrel field patterning, with increases in both the septal area and total barrel field. We provide evidence for functional and structural alterations during brain development induced by in utero exposure to the organophosphorus pesticide chlorpyrifos that may account for the well-established behavioral outcomes. SIGNIFICANCE STATEMENTWe demonstrate persistent alterations to synaptic function and plasticity in the somatosensory cortex following a brief, sub-acute exposure to the organophosphorus pesticide chlorpyrifos in a gestational rat model. These occur in conjunction with structural changes to cellular patterning of this brain region. These previously unknown consequences are potential causal mechanisms to the well-established neurodevelopmental detriments associated with early life exposure to chlorpyrifos. Clarifying these mechanisms could aid in ameliorating or preventing their persistent effects.

neuroscience↗

bioRxiv2025

Stratification along tropical forest succession enhances pollinator diversity via functionally unique canopies

Vertical stratification is a prominent driver of biodiversity in forests. As the proportion of successional forests increases worldwide and canopies are lost at a fast pace, it is crucial to understand the role of stratification as a driver of community recovery and ecological processes during succession, particularly for important plant mutualist groups essential for forest recovery. Within a well-resolved chronosequence in the northwestern Ecuadorian rainforest, we compiled an extensive database of over 20,000 diurnal and nocturnal pollinators and 2,000 interactions with plants and examined the interacting effects of recovery age and stratification on pollinator community and interaction network reassembly. In late successional and old-growth forests, stratification was a stronger predictor of pollinator abundance, alpha-diversity, functional richness and beta-diversity than forest legacy. While most groups were strongly associated with canopies (moths, social bees, and nocturnal bees), orchid bees exhibited an inverse pattern, also presenting the strongest functional response to stratification and recovery status. Within the entire chronosequence (0-38 years of recovery, plus old-growth forests), recovery status and stratification, alongside landscape features, acted synergistically as predictors of pollinator community parameters. Although structurally stable across the chronosequence and between strata, Interaction networks were richer and most distinct in canopies, while the highest pollinator and interaction diversity were found in old-growth canopies. In successional forests, networks were only comparable in size to active disturbances and early recovery with both strata combined, whereas complete networks in old-growth forests were more than twice as large as those in other recovery statuses. Our study underlines the importance of stratification in leveraging pollinator diversity during forest succession. By serving as havens for canopy-adapted species, tropical canopies may thus help safeguard pollination processes in disturbed landscapes.

ecology↗

bioRxiv2024

Integrating natural language processing and genome analysis enables accurate bacterial phenotype prediction

Understanding microbial phenotypes from genomic data is crucial in areas of research including co-evolution, ecology and pathology. This study proposes a new approach to integrate literature-derived information with genomic data to study microbial traits, combining natural language processing (NLP) with functional genome analysis. We applied this methodology to publicly available data to overcome current limitations and provide novel insights into microbial phenotype prediction. We fine-tuned specialized transformer-based large language models to analyze 3.3 million open-access scientific articles, extracting a network of phenotypic information linked to bacterial strains. The network maps relationships between bacterial strains and traits such as pathogenicity, metabolic capacity, and host and biome preference. By functionally annotating reference genome assemblies for strains in the phenotypic network, we were able to predict key genes influencing phenotypes. Our findings align with known phenotypes and reveal novel correlations, leading to the identification of microbial genes relevant in particular disease and host-association phenotypes. The interconnectivity of strains within the network provided further understanding of microbial community interactions, leading to the identification of hub species by inferring trophic connections--insights challenging to extract by means of experimental work. This study demonstrates the potential of machine learning methods to uncover cross-species patterns in microbial gene-phenotype correlations. As the number of sequenced strains and literature descriptions grows exponentially, such methods become crucial for extracting meaningful information and advancing microbiology research.

bioinformatics↗

bioRxiv2024

Large-Scale Quantitative Cross-Linking and Mass Spectrometry Provides New Insight on Protein Conformational Plasticity within Organelles, Cells, and Tissues

Many proteins can exist in multiple conformational states in vivo to achieve distinct functional roles. These states include alternative conformations, variable PTMs, and association with interacting protein, nucleotide, and ligand partners. Quantitative chemical cross-linking of live cells, organelles, or tissues together with mass spectrometry provides the relative abundance of cross-link levels formed in two or more compared samples, which depends both on the relative levels of existent protein conformational states in the compared samples as well as the relative likelihood of the cross-link originating from each. Because cross-link conformational state preferences can vary widely, one expects intra-protein cross-link levels from proteins with high conformational plasticity to display divergent quantitation among samples with differing conformational ensembles. Here we use the large volume of quantitative cross-linking data available on the public XLinkDB database to cluster intra-protein cross-links according to their quantitation in many diverse compared samples to provide the first widescale glimpse of cross-links grouped according to the protein conformational state(s) from which they predominantly originate. We further demonstrate how cluster cross-links can be aligned with any protein structure to assess the likelihood that they were derived from it.

bioinformatics↗

bioRxiv2024

Sex differences in central amygdala glutamate responses to calcitonin gene-related peptide

Women are disproportionately affected by chronic pain compared to men. While societal and environmental factors contribute to this disparity, sex-based biological differences in the processing of pain are also believed to play significant roles. The central lateral nucleus of the amygdala (CeLC) is a key region for the emotional-affective dimension of pain, and a prime target for exploring sex differences in pain processing since a recent study demonstrated sex differences in CGRP actions in this region. Inputs to CeLC from the parabrachial nucleus (PB) play a causal role in aversive processing, and release both glutamate and calcitonin gene-related peptide (CGRP). CGRP is thought to play a crucial role in chronic pain by potentiating glutamatergic signaling in CeLC. However, it is not known if this CGRP-mediated synaptic plasticity occurs similarly in males and females. Here, we tested the hypothesis that female CeLC neurons experience greater potentiation of glutamatergic signaling than males following endogenous CGRP exposure. Using trains of optical stimuli to evoke transient CGRP release from PB terminals in CeLC, we find that subsequent glutamatergic responses are preferentially potentiated in CeLC neurons from female mice. This potentiation was CGRP-dependent and involved a postsynaptic mechanism. This sex difference in CGRP sensitivity may explain sex differences in affective pain processing. Significance statementThe central lateral nucleus of the amygdala (CeLC) receives a dense projection from parabrachial nucleus (PB) neurons that corelease calcitonin gene-related peptide (CGRP) and glutamate following aversive stimuli. This PBCGRP[->]CeLC projection plays a causal role in chronic pain. We show that endogenous CGRP release potentiates glutamate signaling in female, but not male, CeLC neurons. In the context of previous work in male CeLC, this suggests that that females are more sensitive to even transient CGRP release events. Understanding how this sex difference in CGRP sensitivity arises could enhance strategies for treating chronic pain in both women and men.

neuroscience↗

bioRxiv2024

Spatiotemporal transcriptomic niches of complement pathway and serine protease inhibitor activation in aging and infection

Aging is a multifactorial and complex physiological process, affecting every organ with characteristic manifestations. Understanding the molecular mechanisms that drive aging processes is crucial to targeting age-related disorders. Recent reports suggest that severe post-infection syndromes can partially accelerate aging. However, the underlying gene-encoded regulatory interplay, whether being shared or distinct between aging and infection biology are poorly understood. Here, we employed spatial transcriptomics to establish a multi-organ atlas (brain, heart, kidney, liver, lung, and spleen) across the mouse lifespan (4, 17, and 26 months). Dissecting high-quality fresh-frozen tissue samples at unbiased molecular resolution, we found both organ-specific and cross-organ gene dysregulation upon aging. We identified age-related trajectories in gene expression and cell state, some only detectable within their spatial context, and provide validation at subcellular resolution. The most prominent effect was organ-wide immune system activation with spatially variable severity. We therefore evaluated how aging mimics the expression signatures observed in systemic infection, using spatial transcriptomics slices from young mice infected with Plasmodium berghei ANKA. While on the gene level the effect sizes caused by the infection outweighed those of aging, we reveal a shared activation of the early complement pathway (C4b) and serine protease inhibitors (Serpin gene family) within by phenotype distinct spatial niches. We show that this common RNA signature is driven by tissue-specific cell types and eventually affects protein levels in the aged brain, rendering them a target for future mechanistic and drug discovery studies. Taken together, our study provides a coherent in-depth and cross-organ transcriptomics atlas to systematically study aging and infection in the mouse at spatiotemporal resolution. Key highlightsO_LILarge-scale and high-resolution atlas of spatial transcriptomics from six organs to study aging and systemic infection across two mouse cohorts. C_LIO_LIStrong transcriptional alterations found in distinct organ-specific niches for aging and acute malaria, with organ- and cell type-associated immune responses. C_LIO_LIDysregulation of early complement proteases (C4b) and serine protease inhibitors (Serpina3n) as common theme across central nervous system and peripheral organs. C_LI

molecular biology↗

bioRxiv2024

Quantitative design of cell type-specific mRNA stability from microRNA expression data

Limiting expression to target cell types or states is a longstanding goal in gene therapy, which could be met by sensing endogenous microRNA. However, an unclear association between microRNA expression and activity currently hampers such an approach. Here, we probe this relationship by measuring the stability of synthetic microRNA-responsive 3UTRs across 10 cell lines in a library format. By systematically addressing biases in microRNA expression data and confounding factors such as microRNA crosstalk, we demonstrate that a straightforward model can quantitatively predict the stability of reporters containing full microRNA target sites purely from expression data. We use this model to design constructs with previously unattainable response patterns across our cell lines. We then generalize this approach to primary tissues and cells and show that mRNA can be engineered to be differentially active between rested and exhausted T cell states. The rules we derive for microRNA expression data selection and processing should apply to microRNA-responsive devices for any environment with available expression data.

synthetic biology↗

bioRxiv2024

Molecular consequences of SCA5 mutations in the spectrin-repeat domains of beta-III-spectrin

Spinocerebellar ataxia type 5 (SCA5) mutations in the protein {beta}-III-spectrin cluster to the N-terminal actin-binding domain (ABD) and the central spectrin-repeat domains (SRDs). We previously reported that a common molecular consequence of ABD-localized SCA5 mutations is increased actin binding. However, little is known about the molecular consequences of the SRD-localized mutations. It is known that the SRDs of {beta}-spectrin proteins interact with -spectrin to form an /{beta}-spectrin dimer. In addition, it is known that SRDs neighbouring the {beta}-spectrin ABD enhance actin binding. Here, we tested the impact of the SRD-localized R480W and the E532_M544del mutations on the binding of {beta}-III-spectrin to -II-spectrin and actin. Using multiple experimental approaches, we show that both the R480W and E532_M544del mutants can bind -II-spectrin. However, E532_M544del causes partial uncoupling of complementary SRDs in the /{beta}-spectrin dimer. Further, the R480W mutant forms large intracellular inclusions when co-expressed with -II-spectrin in cells, supporting that R480W mutation grossly disrupts the -II/{beta}-III-spectrin physical complex. Moreover, actin-binding assays show that E532_M544del, but not R480W, increases {beta}-III-spectrin actin binding. Altogether, these data support that SRD-localized mutations alter key interactions of {beta}-III-spectrin with -II-spectrin and actin.

biochemistry↗

bioRxiv2024

ERK5 Signaling is Required for Type III IFN-mediated Mucosal Antiviral Responses

Type III interferons (IFN{lambda}) are innate immune cytokines that limit viral replication and coordinate tissue repair through the induction of interferon stimulated genes (ISGs). This response must be tightly regulated to avoid excessive responses that result in the disruption of tissue barrier integrity or inefficient responses that allow for pathogen escape. Here we examine the contribution of Mitogen Activated Protein Kinase (MAPK) signaling on IFN{lambda}-mediated antiviral activity. We find that extracellular-signal-regulated kinase 5 (ERK5), a poorly characterized member of the conventional MAPK family, potentiates the antiviral efficacy of IFN{lambda}. Chemical inhibition and genetic targeting of ERK5 during IFN{lambda} treatment of cells results in a decrease in ISG induction and impaired control of viral infections. This decrease in IFN{lambda} antiviral efficacy in the absence of ERK5 kinase activity corresponded to lowered STAT1 phosphorylation, revealing a noncanonical role for ERK5 in STAT1 activation downstream of IFN{lambda}. In contrast, type I IFN antiviral signaling is largely resistant to ERK5 modulation. Altogether, we identify ERK5 as a potentiator of STAT1 activation, ISG expression, and antiviral activity following type III IFN stimulation. SIGNIFICANCERegulation of type III interferons (IFN{lambda}) at mucosal barriers in response infection to mitigate viral replication and support barrier integrity. The specific mechanistic requirements for MAPK signaling to sustain IFN{lambda}-mediated gene expression have remained elusive. Amongt the least characterized members of the MAPK family, the role of ERK5 in regulating host inflammatory responses has been hampered by off-target effects of kinase inhibitors. Here, we combine pharmacological and genetic approaches to specifically demonstrate that ERK5 promotes antiviral immunity in epithelial cells. Mechanistically, ERK5 enhances the activation of STAT1 in response to IFN stimulation to augment the transcription of IFN-stimulated genes. Our work demonstrates that therapeutic modulation of MAPK and IFN signaling pathway co-integration could distinguish between the protective and deleterious outcomes of IFN expression. One-sentence summaryERK5 potentiates IFN lambda responses.

immunology↗

bioRxiv2024

Perivascular adipose tissue remodeling impairs vasoreactivity in thermoneutral-housed rats

ObjectiveVascular pathology, characterized by impaired vasoreactivity and mitochondrial respiration, differs between the sexes. Housing rats under thermoneutral (TN) conditions causes vascular dysfunction and perturbed metabolism. We hypothesized that perivascular adipose tissue (PVAT), a vasoregulatory adipose depot with brown adipose tissue (BAT) phenotype, remodels to a white adipose (WAT) phenotype in rats housed at TN, driving diminished vasoreactivity in a sex-dependent manner. MethodsMale and female Wistar rats were housed at either room temperature (RT) or TN. Endpoints included changes in PVAT morphology, vasoreactivity in vessels with intact PVAT or transferred to PVAT of the oppositely-housed animal, vessel stiffness, vessel mitochondrial respiration and cellular signaling. ResultsRemodeling of PVAT was observed in rats housed at TN; animals in this environment showed PVAT whitening and displayed diminished aortae vasodilation (p<0.05), different between the sexes. Juxtaposing PVAT from RT rats onto aortae from TN rats in females corrected vasodilation (p<0.05); this did not occur in males. In aortae of all animals housed at TN, mitochondrial respiration was significantly diminished in lipid substrate experiments (p<0.05), and there was significantly less expression of peNOS (p<0.001). ConclusionsThese data are consistent with TN-induced remodeling of PVAT, notably associated with sex-specific blunting of vasoreactivity, diminished mitochondrial respiration, and altered cellular signaling.

physiology↗

bioRxiv2024

Interactome dynamics during heat stress signal transmission and reception

Among evolved molecular mechanisms, cellular stress response to altered environmental conditions to promote survival is among the most fundamental. The presence of stress-induced unfolded or misfolded proteins and molecular registration of these events constitute early steps in cellular stress response. However, what stress-induced changes in protein conformations and protein-protein interactions within cells initiate stress response and how these features are recognized by cellular systems are questions that have remained difficult to answer, requiring new approaches. Quantitative in vivo chemical cross-linking coupled with mass spectrometry (qXL-MS) is an emerging technology that provides new insight on protein conformations, protein-protein interactions and how the interactome changes during perturbation within cells, organelles, and even tissues. In this work, qXL-MS and quantitative proteome analyses were applied to identify significant time-dependent interactome changes that occur prior to large-scale proteome abundance remodeling within cells subjected to heat stress. Interactome changes were identified within minutes of applied heat stress, including stress-induced changes in chaperone systems as expected due to altered functional demand. However, global analysis of all interactome changes revealed the largest significant enrichment in the gene ontology molecular function term of RNA binding. This group included more than 100 proteins among multiple components of protein synthesis machinery, including mRNA binding, spliceosomes, and ribosomes. These interactome data provide new conformational insight on the complex relationship that exists between transcription, translation and cellular stress response mechanisms. Moreover, stress-dependent interactome changes suggest that in addition to conformational stabilization of RNA-binding proteins, adaptation of RNA as interacting ligands offers an additional fitness benefit resultant from generally lower RNA thermal stability. As such, RNA ligands also serve as fundamental temperature sensors that signal stress through decreased conformational regulation of their protein partners as was observed in these interactome dynamics.

molecular biology↗

bioRxiv2024

Hippocampal purinergic P2X7 receptor level is increased in Alzheimer's disease patients, and associated with amyloid and tau pathologies

INTRODUCTIONThe purinergic receptor P2X7R, which is expressed on microglia and astrocytes, plays an important role in Alzheimers disease (AD). We aimed to characterize the alterations in P2X7R expression in AD patients by APOE {varepsilon}4 allele, age and sex, as well as its association with amyloid and tau pathology. METHODSP2X7R staining and quantitative analysis of amyloid, tau, astrocytes and microglia were performed on postmortem hippocampal tissues from 35 AD patients; 31 nondemented controls; caudate/putamen tissue from corticobasal degeneration (CBD), progressive supranuclear palsy (PSP) patients; and bran tissue from aged 3xTg mouse model of AD. RESULTSActivated microglia and reactive astrocytes were observed in the hippocampi of AD patients and exhibited altered morphology with denser cells and pronounced ramifications. Hippocampal P2X7R intensity was greater in the hippocampal subfields of AD patients than in those of nondemented controls and was correlated with amyloid level and Braak stage and was not affected by sex, APOE{varepsilon}4 allele, or age. P2X7R expression increased around A{beta} plaques, cerebral amyloid angiopathy, tau inclusions in the hippocampus from AD patients and tau inclusions in the caudate/putamen from CBD and PSP patients. DISCUSSIONWe found an increased hippocampal P2X7R level in AD compared to non-demented control, which correlated with amyloid and tau pathologies. P2X7R is a potential marker for neuroinflammation in AD.

neuroscience↗

bioRxiv2024

Heterodimerization of Endolysin Isoforms During Bacterial Infection by Staphylococcal Phage {varphi}2638A

AO_SCPLOWBSTRACTC_SCPLOWBacteriophage endolysins targeting Gram-positive bacteria typically feature a modular architecture of one or more enzymatically active domains (EADs) and cell wall binding domains (CBDs). Several endolysins also feature internal translational start sites (iTSSs) that produce short variant (SV) isoforms alongside the full-length (FL) endolysin. While the lytic activity of endolysins and their isoforms has been extensively studied as exogenous agents, the purpose behind producing the SV isoform during the phage infection cycle remains to be explored. In this study, we used staphylococcal phage {varphi}2638A as a model to determine the interplay between its full-length endolysin, Ply2638A, and its SV isoform during phage infection. X-ray crystallography structures and AlphaFold-generated models enabled elucidation of individual functions of the M23 endopeptidase, central amidase, and SH3b domains of Ply2638A. Production of the SV isoform (amidase and SH3b) was confirmed during phage infection and shown to form a heterodimer complex with Ply2638A via inter-amidase domain interactions. Using genetically engineered phage variants, we show that production of both isoforms provides an advantage during phage infection as phages producing only one isoform presented impaired lytic activity, which could be partly restored through recombinant protein complementation of the missing isoform. Importantly, when applied as an antimicrobial protein against Staphylococcus aureus in culture, the activity of Ply2638A remained constant regardless of SV isoform complementation. Drawing from our findings, we propose that SV isoform production provides its biological advantage upon endolysin entry to the periplasmic space to ensure optimal peptidoglycan degradation prior to cell wall lysis and progeny phage release.

molecular biology↗

bioRxiv2023

Bumble bee microbiota shows temporal succession and increase of lactic acid bacteria when exposed to outdoor environments

QuestionThe large earth bumble bee (Bombus terrestris) maintains a social core gut-microbiota, similar as known from the honey bee, which plays an important role for host health and resistance. Experiments under laboratory conditions with commercial hives are limited to these vertically transmitted microbes and neglect variability by environmental influences and external acquisition of microbes. Various environmental and landscape-level factors may have an impact on the gut-microbiota of pollinating insects, with consequences for pollinator health and fitness in agroecosystems. Still, it is not fully clear whether access to a higher vs lower flower diversity will have a significant influence on the bumble bee microbiota. Here, we tested in a semi-field experiment how strongly the bumble bee microbiota changes over time when exposed to different flower diversities within outdoor flight cages. We used commercial hives to distinguish between vertically and horizontally transmitted bacteria, respectively from the nest environment or the exposed outside environment. ResultThe sequential sampling of foraging workers over a period of 35 days indicated a temporal progression of the bumble bee microbiota when exposed to outside conditions. The microbiota became not only more diverse, but changed in composition and variability over time. We observed a major increase in relative abundance of the families Lactobacillaceae, Bifidobacteriaceae and Weeksellaceae. In contrast, major core taxa like Snodgrassella and Gilliamella declined in their relative abundance over time. The genus Lactobacillus showed a high diversity and strain specific turnover, so that only specific ASVs showed an increase over time, while others had a more erratic occurrence pattern. Exposure to different flower diversities had no significant influence on the bumble bee microbiota. ConclusionThe bumble bee microbiota showed a dynamic temporal progression with distinct compositional changes and diversification over time. The exposure of bumble bees to environmental conditions, or environmental microbes, increases dissimilarity and changes the gut-community composition compared to laboratory rearing conditions. This shows the importance of environmental influences on the temporal dynamic and progression of the bumble bee microbiota. Scope statementBumble bees (Bombus terrestris) are, next to the honey bee, commercially important pollinators and widely used to enhance crop pollination service within greenhouse environments. They host a similar, but characteristic, set of core-microbiota which are of known importance for bumble bee health. Despite this, bumble bees harbor their own specific set of symbionts, which do not occur within the honey bee and seem to be more easily influenced by colonization of environmental microbes. While experiments under controlled lab-based rearing conditions often lack the influence of environmental or landscape-level drivers, field-based observation can often not resolve the influence of a single factor. One major unresolved question is which environmental factor influences the microbiota of social pollinators by environmental microbes. Especially whether monocultures (low flower diversity) are per se rather detrimental to microbiota composition compared to more balanced and diverse pollen provisions (high flower diversity). Within this article, we investigated the influence of different flower diversities as potential drivers of the bumble bee gut-microbiota under semi-field conditions. We used outdoor cages which contained a flower diversity gradient to specifically test how a low and high diversity of flower resources could influence the bumble bee microbiota over time.

microbiology↗

bioRxiv2023

DNA methylation profiling identifies TBKBP1 as potent amplifier of cytotoxic activity in CMV-specific human CD8+ T cells

Epigenetic mechanisms stabilize gene expression patterns during CD8+ T cell differentiation. However, although adoptive transfer of virus-specific T cells is clinically applied to reduce the risk of virus infection or reactivation in immunocompromised individuals, the DNA methylation pattern of virus-specific CD8+ T cells is largely unknown. Hence, we here performed whole-genome bisulfite sequencing of cytomegalovirus-specific human CD8+ T cells and found that they display a unique DNA methylation pattern consisting of 79 differentially methylated regions when compared to bulk memory CD8+ T cells. Among them was TBKBP1, coding for TBK-binding protein 1 that can interact with TANK-binding kinase 1 (TBK1) and mediate pro-inflammatory responses in innate immune cells downstream of intracellular virus sensing. Since TBKBP1 has not yet been reported in T cells, we aimed to unravel its role in virus-specific CD8+ T cells. TBKBP1 demethylation in terminal effector CD8+ T cells correlated with TBKBP1 expression and was stable upon long-term in vitro culture. TBKBP1 overexpression resulted in enhanced TBK1 phosphorylation upon stimulation of CD8+ T cells and significantly improved their virus neutralization capacity. Collectively, our data demonstrate that TBKBP1 modulates virus-specific CD8+ T cell responses and could be exploited as therapeutic target to improve adoptive T cell therapies.

immunology↗

bioRxiv2023

Decreased fucosylation impacts epithelial integrity and increases risk for COPD.

COPD causes significant morbidity and mortality worldwide. Epithelial damage is fundamental to disease pathogenesis, although the mechanisms driving disease remain undefined. Published evidence from a COPD cohort (SPIROMICS) and confirmed in a second cohort (COPDgene) demonstrate a polymorphism in Fucosyltransferese-2 (FUT2) is a trans-pQTL for E-cadherin, which is critical in COPD pathogenesis. We found by MALDI-TOF analysis that FUT2 increased terminal fucosylation of E-cadherin. Using atomic force microscopy, we found that FUT2-dependent fucosylation enhanced E-cadherin-E-cadherin bond strength, mediating the improvement in monolayer integrity. Tracheal epithelial cells from Fut2-/- mice have reduced epithelial integrity, which is recovered with reconstitution of Fut2. Overexpression of FUT2 in COPD derived epithelia rescues barrier function. Fut2-/- mice show increased susceptibility in an elastase model of disease developing both emphysema and fibrosis. We propose this is due to the role of FUT2 in proliferation and cell differentiation. Overexpression of FUT2 significantly increased proliferation. Loss of Fut2 results in accumulation of Spc+ cells suggesting a failure of alveolar type 2 cells to undergo transdifferentiation to alveolar type 1. Using a combination of population data, genetically manipulated mouse models, and patient-derived cells, we present a novel mechanism by which post-translational modifications modulate tissue pathology and serve as a proof of concept for the development of a disease-modifying target in COPD.

cell biology↗