Biology subjects

Schlueter, H.

Publications and source records attributed to Schlueter, H..

18 recordsLinked to original sources

bioRxiv2026

The Shewanella oneidensis Fic enzyme SoFic targets the switch-Iregion of EF-Tu for AMPylation

Fic enzymes mediate diverse post-translational modifications, including adenosine monophosphate (AMP) transfer and removal, referred to as AMPylation and deAMPylation, respectively. We identified the prokaryotic translation elongation factor Tu (EF-Tu) as an AMPylation target of the Fic enzyme SoFic. SoFic can constitutively reverse EF-Tu modification via deAMPylation whereas AMPylation depends on SoFic homodimerization. The complex crystal structure between SoFic and EF-Tu confirms a conserved target binding mode across evolutionary distant Fic enzymes. AMPylation disrupts EF-Tus regulatory switch-I region, causing translational inhibition. SoFic furthermore binds to its promotor DNA, suggesting a dual function as transcriptional and translational regulator in bacterial cells. Together, our structural and biochemical data provide valuable insights into the functional and regulatory diversity of Fic enzymes.

biochemistry↗

bioRxiv2026

Temporal Profiling of Upper-Layer Neurons Reveals Changes in the Molecular Landscape Upon Maternal Immune Activation

Neuronal differentiation is a dynamic, multi-layered process that transforms progenitor cells into functionally integrated neurons, yet the coordinated molecular programs underlying this transition remain incompletely understood. Here, we define the developmental trajectory of murine upper-layer cortical neurons using an integrated multi-omics approach combining transcriptomics and proteomics across key stages of neurogenesis. We find that neuronal identity emerges gradually through coordinated transitions from RNA processing and splicing programs toward synaptic and metabolic maturation. Integration of matched transcriptomic and proteomic datasets reveals a compact set of concordant molecular modules that define this trajectory, highlighting post-transcriptional regulation as a central driver of neuronal maturation. We further show that maternal immune activation (MIA), a model of prenatal inflammation, deranges this developmental program. MIA induces sustained upregulation of Wnt signalling pathways alongside a downregulation of synaptic regulators, without detectable global alterations in DNA methylation. These molecular changes are accompanied by defects in neuronal positioning during cortical development, linking altered molecular trajectories to functional outcomes. Together, our findings establish a temporal molecular framework of neuronal differentiation and demonstrate that prenatal environmental perturbations reshape cortical development primarily through post-transcriptional and signalling-based mechanisms.

neuroscience↗

bioRxiv2026

Integrated kidney and urine proteomics define encrypted antimicrobial peptides as effectors of host defence in human pyelonephritis

Antimicrobial peptides (AMPs) are key effectors of host defence, however, their functional deployment across renal tissue and urine in pyelonephritis (PN) remains incompletely understood. Here, we integrate kidney and urine proteomics with urinary peptidomics and computational prediction to define AMP organisation and function. Proteomic analysis indicated coordinated induction of multiple AMPs in infected kidneys. These patterns were recapitulated in the urinary proteome, where AMP abundance correlated with leukocyte counts. Multiplex immunofluorescence microscopy localised these AMPs to myeloid cells, identifying them as central effector sources. Importantly, analysis of the urine peptidome revealed multiple encrypted AMPs (EPs), which arise from proteolytic processing of precursor proteins. To systematically assess their relevance for host defence, we applied an ensemble of machine learning-based predictors to prioritise candidates with activity in the urinary environment. This approach identified several potential EPs, among which the S100A12-derived peptide Calcitermin was confirmed in patient urine. Furthermore, it exerts antibacterial activity against uropathogenic E. coli (UPEC) and modulates myeloid cell responses. Together, these findings define a coordinated and compartmentalised AMP defence programme in human PN that extends beyond increased peptide expression, highlighting EPs as functionally relevant effectors with therapeutic potential.

immunology↗

bioRxiv2026

Lactate-dependent metabolic rewiring is associated with CD163+ macrophage polarization in human pyelonephritis

Chronic pyelonephritis is characterized by persistent bacterial infection of the kidney and a dysregulated immune response that promotes disease progression. Macrophages are central regulators of antibacterial immunity in the urinary tract. However, the mechanisms underlying their functional reprogramming in chronic infection remain poorly understood. Here, we identify lactate as a key metabolic determinant of macrophage polarization in human pyelonephritis. Proteomic analysis of human kidney tissue revealed extensive metabolic remodelling, including upregulation of enzymes involved in glycolysis. Consistent with this, lactate levels were significantly elevated in urine and plasma of pyelonephritis patients. Concomitantly, we observed a pronounced accumulation of CD163 macrophages in infected kidneys, representing a distinct macrophage subset with immunomodulatory function. Correlation-based network analysis revealed a strong association between CD163 and lactate dehydrogenase A, supporting a functional association between lactate metabolism and macrophage polarization. Mechanistically, exposure of murine bone marrow-derived macrophages to lactate induced intracellular protein lactylation and promoted polarization toward a CD163 phenotype, defining a metabolically imprinted macrophage state distinct from classical activation paradigms. Proteomic profiling demonstrated extensive remodelling of macrophage protein expression in response to lactate with significant alteration of mediators of phagocytosis, Toll-like receptor signalling, and interferon response. Together, these findings identify lactate as a potent metabolic driver of macrophage reprogramming and establish a foundation for investigating lactylation-dependent immune dysfunction in chronic pyelonephritis.

immunology↗

bioRxiv2026

Histone neutralization protects the ischemic brain against stroke-associated pneumonia

Bacterial pneumonia aggravates ischemic stroke via mechanisms that still remain to be determined. In ischemic stroke patients and mice exposed to middle cerebral artery occlusion, we show that stroke-associated pneumonia markedly worsens clinical stroke outcome. In mice, pneumonia induced 3 days after stroke impaired neurological recovery and increased brain neutrophil infiltrates, blood-brain barrier breakdown, cerebral microvascular thrombosis, and progressive brain atrophy. The antibiotic amoxicillin only partially ameliorated pneumonia-associated neurological deficits and neutrophil infiltrates. Neutrophils were critical mediators of pneumonia-induced blood-brain barrier breakdown and microvascular thrombosis. Notably, administration of a neutralizing anti-histone antibody during pneumonia--unlike degradation or blockade of neutrophil extracellular trap formation or myeloperoxidase inhibition--restored long-term neurological recovery and prevented brain atrophy in stroke-associated pneumonia mice. This study identifies extracellular histones as key drivers of secondary inflammatory brain injury and establishes histone neutralization as a therapeutic strategy with an extended treatment window in the post-acute stroke phase. One Sentence SummaryNeutralizing extracellular histones reverses pneumonia-driven secondary brain injury and restores long-term recovery after ischemic stroke.

neuroscience↗

bioRxiv2025

TSP1/TGF-β1 drives arachidonic acid metabolism to orchestrate neutrophil swarming

Neutrophil swarming has emerged as a conserved multicellular behaviour observed across tissues and pathological contexts. Yet, the molecular cues and the spatially coordinated cellular circuits that drive the process of neutrophil swarming leading to cluster formation remain poorly understood. Here, we combine spatial proteomics and lipid profiling in a model of urinary tract infection to define the epithelial-immune circuits driving neutrophil cluster formation. We identify thrombospondin-1 (TSP1)-mediated activation of transforming growth factor beta 1 (TGF-{beta}1) as key epithelial signal licensing neutrophil clustering and enhancing bacterial control. Spatial lipid analysis further reveals that TSP1/TGF-{beta}1 signalling locally activates arachidonic acid metabolism in epithelial neutrophils, with 5-lipoxygenase dependent leukotriene synthesis required for swarm formation and infection clearance. These findings uncover a spatially coordinated defence mechanism in which epithelial-derived TSP1/TGF-{beta}1 engages neutrophil lipid metabolism to orchestrate neutrophil swarming behaviour and reinforce antibacterial immunity. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/688397v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@114ea92org.highwire.dtl.DTLVardef@345244org.highwire.dtl.DTLVardef@10520baorg.highwire.dtl.DTLVardef@1a7c0fe_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG

immunology↗

bioRxiv2025

Infrared Laser Sampling of Low Volumes Reveals Marker Lipids in Palatine Tonsil Carcinoma via Shotgun Lipidomics

Complete surgical resection is essential for oropharyngeal squamous cell carcinoma (OPSCC) therapy, underscoring the need for improved intraoperative margin assessment. To advance in-vivo diagnostics of OPSCC, Nanosecond infrared laser (NIRL) tissue sampling combined with shotgun lipidomic analysis reveals lipidome differences between OPSCC tissue and adjacent healthy tissue. Ablations were performed on tonsil squamous cell carcinoma on n=28 samples from 11 patients with an established chamber setup and a subset of n=6 samples from three patients with a custom-made handheld applicator. Welchs t-test results (p=0.05, two-fold change) revealed a similar OPSCC lipid profile in 7 of 11 patients. Potential tumor lipid markers were identified as consistently significantly increased, despite biological heterogeneity of the samples, underscoring their potential diagnostic value. Tissue ablation with a custom-made handheld applicator coupled to a laser fiber was successful and the lipidomic analysis was consistent to the chamber setup. Although our setup is currently limited by an ablation time exceeding one minute, this study demonstrates the potential of the handheld applicator for future applications such as endoscopy or intraoperative diagnostics.

cancer biology↗

bioRxiv2025

Extracellular vesicles released from cortical neurons influence spontaneous activity of recipient neurons.

Extracellular vesicles (EVs) are membranous structures that cells release into the extracellular space. EVs carry various molecules such as proteins, lipids, and nucleic acids, and serve as specialized transporters to influence other cells. In the central nervous system, EVs have been linked to many important processes, including intercellular communication, but molecular details of their physiological functions are not fully understood. Our study aimed to investigate how EVs are released by neuronal cells, and how they affect the neuronal activity of other recipient neurons. We show that mature primary cortical neurons release EVs from both their soma and dendrites. EVs released from neurons closely resemble non-neuronal EVs regarding size and marker proteins and proteomic analyses showed that neuronally released EVs contain proteins typically acting in pre- and post-synaptic compartments. Interestingly, our analysis revealed that EVs alter spontaneous activity in target neurons by increasing the amplitude of postsynaptic potentials. In summary, our findings elaborates on the role of EVs in synaptic activity modulation in neurons mediated by glutamate receptors.

neuroscience↗

bioRxiv2024

PTX3 Governs Fibroblast-Epithelial Dynamics in Lung Injury and Repair

Dysfunctional interactions between fibroblasts and epithelial cells contribute to the progression of chronic lung diseases, including idiopathic pulmonary fibrosis (IPF). In this study, we utilized a coculture model of human small airway epithelial cells and fibroblasts to investigate intercellular communication during disease progression. Our transcriptomic and proteomic profiling reveal that fibroblasts repair epithelial cells in acute injury by boosting epithelial fatty acid metabolism; conversely, they exacerbate epithelial damage in chronic injury scenarios. By delineating regulators involved in these responses, we identified pentraxin 3 (PTX3) as a key antifibrotic factor secreted by fibroblasts in response to acute epithelial injury. Importantly, PTX3 levels are decreased in bronchoalveolar lavage (BAL) samples from IPF patients compared to non-fibrotic controls, indicating a potential link between diminished PTX3 levels and fibrosis progression. Furthermore, adding PTX3 to chronically injured epithelial-fibroblast cocultures mitigated the pro-fibrotic response and restored the epithelial barrier integrity. These findings highlight the dual roles of fibroblasts and the critical function of PTX3 in lung injury and repair, offering insights for therapeutic strategies.

molecular biology↗

bioRxiv2024

Co-activation of LIN28A and CTNNB1 disturbs cortical neuronal migration and pia mater integrity

Developmental signalling pathways act in stage and tissue dependent relation and mis-activation can drive tumour formation. The RNA-binding protein LIN28A maintains stemness and is overexpressed in embryonal brain tumours. Activating mutations of CTNNB1 - the WNT pathway effector - have been reported in respective brain tumours. The aim of this study was to investigate the interplay of these oncogenic proteins during embryonal brain development. The combination of both oncogenic factors did not lead to brain tumour formation but resulted in disturbed lamination and impaired cell migration in the cerebral cortex. Spatially resolved proteome analysis revealed imbalances of the extracellular matrix protein LAMB1 and its receptors RPSA and ITGB1 accompanied by a porous pial border and overmigration of neural cells. Cajal-Retzius cells were misplaced in deeper cortex regions without affecting general REELIN levels and additional reduced levels of -DYSTROGLYCAN. Taken together, the interplay of LIN28A and CTNNB1 resulted in a cortical migration disorder showing histomorphological and molecular similarities to human Cobblestone lissencephaly (type 2), highlighting novel implications of the oncogene LIN28A in extracellular matrix integrity.

neuroscience↗

bioRxiv2024

Efficient enzyme-free isolation of brain-derived extracellular vesicles

Extracellular vesicles (EVs) have gained significant attention as pathology mediators and potential diagnostic tools for neurodegenerative diseases. However, isolation of brain-derived EVs (BDEVs) from tissue remains challenging, often involving enzymatic digestion steps that may compromise the integrity of EV proteins and overall functionality. Here, we describe that collagenase digestion, commonly used for BDEV isolation, produces undesired protein cleavage of EV-associated proteins in brain tissue homogenates and cell-derived EVs. In order to avoid this effect, we studied the possibility of isolating BDEVs with a reduced amount of collagenase or without any protease. Characterization of the isolated BDEVs revealed their characteristic morphology and size distribution with both approaches. However, we revealed that even minor enzymatic digestion induces artificial proteolytic processing in key BDEV markers, such as Flotillin-1, CD81, and the cellular prion protein (PrPC), whereas avoiding enzymatic treatment completely preserves their integrity. We found no differences in mRNA and protein content between non-enzymatically and enzymatically isolated BDEVs, suggesting that we are purifying the same BDEV populations with both approaches. Intriguingly, the lack of Golgi marker GM130 signal, often referred to as contamination contamination-negative marker in EV preparations, seems to result from enzymatic digestion rather than from its actual absence in BDEV samples. Overall, we show that non-enzymatic isolation of EVs from brain tissue is possible and avoids artificial pruning of proteins while achieving a high BDEV yield and purity. This protocol will help to understand the functions of BDEV in a near-physiological setting, thus opening new research approaches.

neuroscience↗

bioRxiv2023

Spatial Transcriptomic Characterization of Novel Pathologic Niches in IPF

An unmet medical need persists in Idiopathic Pulmonary fibrosis (IPF), for which treatments additional to anti-fibrotic therapy are needed. Single cell RNA sequencing (scRNA-seq) has advanced our understanding of IPF with cell type-specific insights but lacks cellular tissue context. Spatial transcriptomics addresses this by providing spatially resolved gene expression, enabling gene and cell type localization within the tissue environment. We profiled IPF and control patient lung tissue sections using spatial transcriptomics and combined the data with an atlas of integrated IPF scRNA-seq datasets. Through computational analysis, we identified three disease-associated pathologic niches with unique cellular composition / localization and analyzed their cell-cell communication. We identified the Fibrotic niche, comprising Myofibroblasts and Aberrant Basaloid cells, preferentially located around airways and close to the Airway Macrophage niche in the lumen, containing SPP1+ Macrophages. We also identified the Immune niche, distinct foci of lymphoid cells in fibrotic tissue, surrounded by remodeled endothelial vessels. TEASERSpatial transcriptomics localizes genes and cell types in the tissue and identifies pathological cellular niches in IPF and control lungs.

systems biology↗

bioRxiv2023

Cystatin C loaded in brain-derived extracellular vesicles rescues synapses after ischemic insult in vitro and in vivo

Synaptic loss is an early event in the undersupplied but not yet irreversibly injured penumbra area after an ischemic stroke. Promoting synaptic preservation in this area would likely improve functional neurological recovery. In the present study, we aimed to detect proteins involved in endogenous protection mechanisms of synapses in the penumbra after stroke and to analyse the potential beneficial effect of these candidates for a prospective stroke treatment. For this, we performed Liquid Chromatography coupled to Mass Spectrometry (LC-MS)-based proteomics of synaptosomes isolated from the ipsilateral hemispheres of mice subjected to experimental stroke at different time points (24 h, 4 and 7 days) and compared them to sham-operated mice. Proteomic analyses indicated that among the differentially expressed proteins between the two groups, cystatin C (CysC) was significantly increased at 24 h and 4 days following stroke, before returning to steady-state levels at 7 days, thus indicating a potential transient and intrinsic rescue mechanism attempt of neurons. When CysC was applied to primary neuronal cultures subjected to an in vitro model of ischemic damage, this treatment significantly improved the preservation of synaptic structures. Notably, similar effects were observed when CysC was loaded into brain-derived extracellular vesicles (BDEVs). Finally, when CysC contained in BDEVs was administered intracerebroventricularly to stroked mice, it significantly increased the expression of synaptic markers such as SNAP25, Homer-1, and NCAM in the penumbra area compared to the group supplied with empty BDEVs. Thus, we show that CysC-loaded BDEVs promote synaptic protection after ischemic damage in vitro and in vivo, opening the possibility of a therapeutic use in stroke patients.

neuroscience↗

bioRxiv2023

Multiomic profiling of medulloblastoma reveals subtype-specific targetable alterations at the proteome and N-glycan level

Medulloblastomas (MBs) are malignant pediatric brain tumors that are molecularly and clinically very heterogenous. To unravel phenotypically relevant MB subtypes, we compiled a harmonized proteome dataset of 167 MBs and integrated findings with DNA methylation and N-glycome data. Six proteome MB subtypes emerged, that could be assigned to two main molecular programs: transcription/translation (pSHHt, pWNT and pGroup3-Myc), and synapses/immunological processes (pSHHs, pGroup3 and pGroup4). Multiomic analysis revealed different conservation levels of proteome features across MB subtypes at the DNA-methylation level. Aggressive pGroup3-Myc MBs and favorable pWNT MBs were most similar in cluster hierarchies concerning overall proteome patterns but showed different protein abundances of the vincristine resistance associated multiprotein complex TriC/CCT and of N-glycan turnover associated factors. The N-glycome reflected proteome subtypes and complex-bisecting N-glycans characterized pGroup3-Myc tumors. Our results shed light on new targetable alterations in MB and set a foundation for potential immunotherapies targeting glycan structures. SignificanceWhereas the application of omics technologies has significantly improved MB tumor classification and treatment stratification, it is still of debate, which features predict best clinical outcome. Moreover, treatment options - especially for high-risk groups - are still unsatisfactory. In contrast to nucleic acids, the proteome and their N-glycans may reflect the phenotype of a tumor in a more direct way and thus hold the potential to discover clinically relevant phenotypes and potentially targetable pathways. We show that these analyses are feasible on formalin fixed and paraffine embedded tissue. Compiling a comprehensive MB dataset, we detected new biomarkers and characteristics for high- and low-risk MB subtypes that were not reflected by other omic data modalities before. Specifically, we identified subtype specific abundance differences in proteins of the vincristine resistance associated multiprotein complex TriC/CCT and in proteins involved in N-glycan turnover. Changes in the N-glycans are considered as potential hallmarks of cancer and we show that N-glycan profiles can distinguish MB subtypes. These tumor-specific N-glycan structures hold a strong potential as new biomarkers, as well as immunotherapy targets. Highlights- Integration of in-house proteome data on formalin fixated paraffine embedded medulloblastoma (MB) and publicly available datasets enables large scale proteome analysis of MB - Six proteome MB subtypes can be assigned to two main molecular programs: replication/ translation versus synapse/immune system - Identification and validation of IHC compatible protein-biomarkers for high and low risk MB subtypes, such as TNC and PALMD. - Subtype specific correlation of the DNA methylome and the proteome reveals different conserved molecular characteristics across MB subtypes. - pGroup3-Myc subtype MBs are associated with high-risk features including high abundances of vincristine resistance associated TriC/CCT member proteins - Proteome MB subtypes show differential N-glycosylation patterns, revealing complex-bisecting glycans as potentially immunotargetable hallmarks of the high risk pGroup3-Myc subtype.

cancer biology↗

bioRxiv2020

Defective NETs Clearance contributes to sustained FXII Activation in COVID-19-associated Pulmonary Thrombo-Inflammation

BackgroundCoagulopathy and inflammation are hallmarks of Coronavirus disease 2019 (COVID-19) and are associated with increased mortality. Clinical and experimental data have revealed a role for neutrophil extracellular traps (NETs) in COVID-19 disease. The mechanisms that drive thrombo-inflammation in COVID-19 are poorly understood. MethodsWe performed proteomic analysis and immunostaining of postmortem lung tissues from COVID-19 patients and patients with other lung pathologies. We further compared coagulation factor XII (FXII) and DNase activities in plasma samples from COVID-19 patients and healthy control donors and determined NET-induced Factor XIII (FXII) activation using a chromogenic substrate assay. FindingsFXII expression and activity were increased in the lung parenchyma, within the pulmonary vasculature and in fibrin-rich alveolar spaces of postmortem lung tissues from COVID-19 patients. In agreement with this, plasma FXII activation (FXIIa) was increased in samples from COVID-19 patients. Furthermore, FXIIa colocalized with NETs in COVID-19 lung tissue indicating that NETs accumulation leads to FXII contact activation in COVID-19. We further showed that an accumulation of NETs is partially due to impaired NET clearance by extracellular DNases as DNase substitution improved NET dissolution and reduced FXII activation in vitro. InterpretationCollectively, our study supports that the NETs/FXII axis contributes to the pathogenic chain of procoagulant and proinflammatory responses in COVID-19. Targeting both, NETs and FXIIa, could provide a strategy to mitigate COVID-19-induced thrombo-inflammation. FundingThis study was supported by the European Union (840189), the Werner Otto Medical Foundation Hamburg (8/95) and the German Research Foundation (FR4239/1-1, A11/SFB877, B08/SFB841 and P06/KFO306).

molecular biology↗

bioRxiv2020

N-terminal VP1 truncations favor T=1 norovirus-like particles

Noroviruses cause immense sporadic gastroenteritis outbreaks worldwide. Emerging genotypes, which are divided based on VP1 sequence, further enhance this public threat. Self-assembling properties of the human norovirus major capsid protein VP1 are crucial for using virus-like particles (VLPs) for vaccine development. However, there is no vaccine available yet. Here, VLPs from different variants produced in insect cells are characterized in detail using a set of biophysical and structural tools. We are using native mass spectrometry, gas-phase electrophoretic mobility molecular analysis and proteomics to get clear insights into particle size, structure, composition as well as stability. Generally, noroviruses have been known to form mainly T=3 particles. Importantly, we identify a major truncation in the capsid proteins as a likely cause for the formation of merely T=1 particles. For vaccine development, particle production needs to be a reproducible, reliable process. Understanding the underlying processes in capsid size variation will help to produce particles of a defined capsid size presenting antigens consistent with intact virions. Next to vaccine production itself, this would be immensely beneficial for bio-/nano-technological approaches using viral particles as carriers or triggers for immunological reactions.

biophysics↗

bioRxiv2019

Flagella methylation promotes bacterial adhesion and host cell invasion

The flagellum is the motility device of many bacteria and the long external filament is made of several thousand copies of a single protein, flagellin. While posttranslational modifications of flagellin are common among bacterial pathogens, the role of lysine methylation remained unknown. Here, we show that both flagellins of Salmonella enterica, FliC and FljB, are methylated at surface-exposed lysine residues. A Salmonella mutant deficient in flagellin methylation was outcompeted for gut colonization in a gastroenteritis mouse model. In support, methylation of flagellin promoted invasion of epithelial cells in vitro. Lysine methylation increased the surface hydrophobicity of flagellin and enhanced flagella-dependent adhesion of Salmonella to phosphatidylcholine vesicles and epithelial cells. In summary, posttranslational flagellin methylation constitutes a novel mechanism how flagellated bacteria facilitate adhesion to hydrophobic host cell surfaces and thereby contributes to efficient gut colonization and successful infection of the host.

microbiology↗

bioRxiv2019

FLASHDeconv: Ultrafast, high-quality feature deconvolution for top-down proteomics

Feature deconvolution, the determination of intact proteoform masses, is crucial for top-down proteomics, but currently suffers from long runtimes and quality issues. We present FLASHDeconv, an algorithm based on a simple transformation of mass spectra, which turns deconvolution into the search for constant patterns thus greatly accelerating the process. We show higher deconvolution quality and two to three orders of magnitude faster execution speed than existing approaches.

bioinformatics↗