Biology subjects

Krueger, T.

Publications and source records attributed to Krueger, T..

31 records · Page 2Linked to original sources

bioRxiv2022

Red blood cell lingering modulates hematocrit distribution in the microcirculation

The distribution of red blood cells (RBCs) in the microcirculation determines the oxygen delivery and solute transport to tissues. This process relies on the partitioning of RBCs at successive bifurcations throughout the microvascular network and it is known since the last century that RBCs partition disproportionately to the fractional blood flow rate, therefore leading to heterogeneity of the hematocrit (i.e. volume fraction of RBCs in blood) in microvessels. Usually, downstream of a microvascular bifurcation, the vessel branch with a higher fraction of blood flow receives an even higher fraction of RBC flux. However, both temporal and time-average deviations from this phaseseparation law have been observed in recent works. Here, we quantify how the microscopic behavior of RBCs lingering (i.e. RBCs temporarily residing near the bifurcation apex with diminished velocity) influences their partitioning, through combined in vivo experiments and in silico simulations. We developed an approach to quantify the cell lingering at highly-confined capillary-level bifurcations and demonstrate that it correlates with deviations of the phase-separation process from established empirical predictions by Pries et al. Furthermore, we shed light on how the bifurcation geometry and cell membrane rigidity can affect the lingering behavior of RBCs, e.g. rigid cells tend to linger less than softer ones. Taken together, RBC lingering is an important mechanism that should be considered when studying how abnormal RBC rigidity in diseases such as malaria and sickle-cell disease could hinder the microcirculatory blood flow or how the vascular networks are altered under pathological conditions (e.g. thrombosis, aneurysm).

biophysics↗

bioRxiv2022

Functional characterization of the RNA interference pathway in A. fumigatus reveals its potential for antifungal therapy

The RNA interference (RNAi) pathway has evolved numerous functionalities in eukaryotes, with many on display in Kingdom Fungi. RNAi can regulate gene expression, facilitate drug resistance, or even be altogether lost to improve growth potential in some fungal pathogens. In the WHO fungal priority pathogen, Aspergillus fumigatus, the RNAi system is known to be intact and functional. To extend our limited understanding of A. fumigatus RNAi, we first investigated the genetic variation in RNAi-associated genes in a collection of 217 environmental and 83 clinical genomes, where we found that RNAi components are conserved even in clinical strains. Using endogenously expressed inverted-repeat transgenes complementary to a conditionally essential gene (pabA) or a nonessential gene (pksP), we determined that a subset of the RNAi componentry is active in inverted-repeat transgene silencing in conidia and mycelium. Analysis of mRNA-seq data from RNAi double-knockout strains linked the A. fumigatus dicer-like enzymes (DclA/B) and RNA-dependent RNA polymerases (RrpA/B) to regulation of conidial ribosome biogenesis genes; however, surprisingly few endogenous small RNAs were identified in conidia that could explain this broad change. Although RNAi was not clearly linked to growth or stress response defects in the RNAi knockouts, serial passaging of RNAi knockout strains for six generations resulted in lineages with diminished spore production over time, indicating that loss of RNAi can exert a fitness cost on the fungus. Cumulatively, A. fumigatus RNAi appears to play an active role in defense against double-stranded RNA species alongside a previously unappreciated housekeeping function in regulation of conidial ribosomal biogenesis genes.

microbiology↗

bioRxiv2022

Red blood cell dynamics in extravascular biological tissues modelled as canonical disordered porous media

The dynamics of blood flow in the smallest vessels and passages of the human body, where the cellular character of blood becomes prominent, plays a dominant role in the transport and exchange of solutes. Recent studies have revealed that the micro-haemodynamics of a vascular network is underpinned by its interconnected structure, and certain structural alterations such as capillary dilation and blockage can substantially change blood flow patterns. However, for extravascular media with disordered microstructure (e.g., the porous intervillous space in the placenta), it remains unclear how the mediums structure affects the haemodynamics. Here, we simulate cellular blood flow in simple models of canonical porous media representative of extravascular biological tissue, with corroborative microfluidic experiments performed for validation purposes. For the media considered here, we observe three main effects: first, the relative apparent viscosity of blood increases with the structural disorder of the medium; second, the presence of red blood cells (RBCs) dynamically alters the flow distribution in the medium; third, increased structural disorder of the medium can promote a more homogeneous distribution of RBCs. Our findings contribute to a better understanding of the cellscale haemodynamics that mediates the relationship linking the function of certain biological tissues to their microstructure.

biophysics↗

bioRxiv2022

Re-direction of phagosomes to the recycling expulsion pathway by a fungal pathogen

The analysis of host-pathogen interactions bears the potential to discover novel pathogenicity mechanisms and to obtain novel insights into basic mechanisms of cell biology. Here, we obtained unprecedented insight into both. We discovered that the HscA protein on the conidial surface of the clinically important human-pathogenic fungus Aspergillus fumigatus acts as an effector protein. It inhibits phagosome maturation and reprograms phagosomes for expulsion of conidia. HscA anchors the human p11 protein to phagosomes. p11 is a decisive factor for targeting phagosomes either to the degradative or secretory pathway. The relevance of our findings is indicated by the identification of an SNP in the non-coding region of the human p11 gene that affects its translation and is associated with heightened susceptibility to invasive pulmonary aspergillosis.

cell biology↗

bioRxiv2022

Ubiquitous bacterial polyketides induce cross-kingdom microbial interactions

Although the interaction of prokaryotic and eukaryotic microorganisms is critical for the functioning of ecosystems, knowledge of the processes driving microbial interactions within communities is in its infancy. We previously reported that the soil bacterium Streptomyces iranensis specifically triggers the production of natural products in the fungus Aspergillus nidulans. Here, we discovered that arginine-derived polyketides serve as the bacterial signals for this induction. Arginine-derived polyketide-producing bacteria occur world wide. These producer bacteria and the fungi that decode and respond to this signal can be co-isolated from the same soil sample. Arginine-derived polyketides impact surrounding microorganisms both directly as well as indirectly, by inducing the production of natural products in fungi that further influence the composition of microbial consortia. One-Sentence SummaryUbiquitous bacterial polyketides are universal components of the chemical network for microbial communication

microbiology↗

bioRxiv2020

Vessel compression biases red blood cell partitioning at bifurcations in a haematocrit-dependent manner: implications for tumour blood flow

The tumour microenvironment is abnormal and associated with tumour tissue hypoxia, immunosuppression, and poor response to treatment. One important abnormality present in tumours is vessel compression. Vessel decompression has been shown to increase survival rates in animal models via enhanced and more homogeneous oxygenation. However, our knowledge of the biophysical mechanisms linking tumour decompression to improved tumour oxygenation is limited. In this study, we propose a computational model to investigate the impact of vessel compression on red blood cell (RBC) dynamics in tumour vascular networks. Our results demonstrate that vessel compression can alter RBC partitioning at bifurcations in a haematocrit-dependent and flowrate-independent manner. We identify RBC focussing due to cross-streamline migration as the mechanism responsible and characterise the spatiotemporal recovery dynamics controlling downstream partitioning. Based on this knowledge, we formulate a reduced-order model that will help future research to elucidate how these effects propagate at a whole vascular network level. These findings contribute to the mechanistic understanding of haemodilution in tumour vascular networks and oxygen homogenisation following pharmacological solid tumour decompression.

cancer biology↗

bioRxiv2020

Biotinylated surfome profiling identifies potential biomarkers for diagnosis and therapy of Aspergillus fumigatus infection

Aspergillus fumigatus is one of the most common airborne fungi capable of causing invasive mycoses in immunocompromised patients and allergic diseases in susceptible individuals. In both cases, fungal surface proteins mediate the first contact with the human immune system to evade immune responses or to induce hypersensitivity. Several methods have been established to study the surface proteome (surfome) of A. fumigatus, like trypsin shaving, glucanase treatment, or formic acid extraction. Biotinylation coupled with LC-MS/MS identification of peptides is a particularly efficient method to identify the surface exposed regions of proteins that potentially mediate interaction with the host. After biotinylation of surface proteins during spore germination, we detected 314 different surface proteins, including several well-known proteins like RodA, CcpA, and DppV, as well as several allergens, heat shock proteins (HSPs), and previously undescribed surface proteins. Using immunofluorescence microscopy, we confirmed the surface localization of three HSPs, which may have moonlighting functions. Collectively, our study generated a comprehensive data set of the A. fumigatus surfome, which complements already existing A. fumigatus surface proteome data and allows us to propose a common core set of A. fumigatus surface proteins. In addition, our study uncovers the surface-exposed regions of many proteins on the surface of spores or hyphae. These surface exposed regions are candidates for direct interaction with host cells and may represent antigenic epitopes that either induce protective immune responses or mediate immune evasion. Thus, the comprehensive datasets provided and compiled here represent reasonable immunotherapy and diagnostic targets for future investigations. HIGHLIGHTSO_LISurface protein biotinylation coupled with LC-MS/MS analysis provides a comprehensive dataset of the A. fumigatus surface proteome. C_LIO_LI314 different A. fumigatus proteins (including immunoreactive proteins, and virulence factors) with surface exposed regions were detected. C_LIO_LISurface localization of three Hsp70 chaperones was confirmed by protein tagging coupled with immunofluorescence. C_LIO_LIBy comparison with other surfome datasets, a core surfome of A. fumigatus was defined, which provides possible biomarkers for diagnosis or therapy. C_LI SIGNIFICANCEAspergillus fumigatus is the most important airborne human pathogenic mold, capable of causing both life-threatening invasive pulmonary aspergillosis in immunocompromised patients and allergic infections in atopic individuals. Despite its obvious medical relevance, timely diagnosis and efficient antifungal treatment of A. fumigatus infection remains a major challenge. Proteins on the surface of conidia (asexually produced spores) and mycelium directly mediate host-pathogen interaction and also may serve as targets for diagnosis and immunotherapy. However, the similarity of protein sequences between A. fumigatus and other organisms, and sometimes even the human host, makes selection of targets for immunological-based studies difficult. Here, using surface protein biotinylation coupled with LC-MS/MS analysis, we identified hundreds of A. fumigatus surface proteins with exposed regions, further defining putative targets for possible diagnostic and immunotherapeutic design.

microbiology↗

bioRxiv2020

The dynamic surface proteomes of allergenic fungal conidia

Fungal spores and hyphal fragments play an important role as allergens in respiratory diseases. In this study, we performed trypsin shaving and secretome analyses to identify the surface-exposed proteins and secreted/shed proteins of Aspergillus fumigatus conidia, respectively. We investigated the surface proteome under different conditions, including temperature variation and germination. We found that the surface proteome of resting A. fumigatus conidia is not static, but instead unexpectedly dynamic, as evidenced by drastically different surface proteomes under different growth conditions. Knockouts of two abundant A. fumigatus surface proteins, ScwA and CweA, were found to function only in fine-tuning the cell wall stress response, implying that the conidial surface is very robust against perturbations. We then compared the surface proteome of A. fumigatus to other allergy-inducing molds, including Alternaria alternata, Penicillium rubens, and Cladosporium herbarum, and performed comparative proteomics on resting and swollen conidia, as well as secreted proteins from germinating conidia. We detected 125 protein ortholog groups, including 80 with putative catalytic activity, in the extracellular region of all four molds, and 42 nonorthologous proteins produced solely by A. fumigatus. Ultimately, this study highlights the dynamic nature of the A. fumigatus conidial surface and provides targets for future diagnostics and immunotherapy.

microbiology↗

bioRxiv2019

Abnormal morphology biases haematocrit distribution in tumour vasculature and contributes to heterogeneity in tissue oxygenation

Oxygen heterogeneity in solid tumours is recognised as a limiting factor for therapeutic efficacy. This heterogeneity arises from the abnormal vascular structure of the tumour, but the precise mechanisms linking abnormal structure and compromised oxygen transport are only partially understood. In this paper, we investigate the role that RBC transport plays in establishing oxygen heterogeneity in tumour tissue. We focus on heterogeneity driven by network effects, which are challenging to observe experimentally due to the reduced fields of view typically considered. Motivated by our findings of abnormal vascular patterns linked to deviations from current RBC transport theory, we calculate average vessel lengths [Formula] and diameters [Formula] from tumour allografts of three cancer cell lines and observe a substantial reduction in the ratio [Formula] compared to physiological conditions. Mathematical modelling reveals that small values of the ratio{lambda} (i.e. {lambda} < 6) can bias haematocrit distribution in tumour vascular networks and drive heterogeneous oxygenation of tumour tissue. Finally, we show an increase in the value of{lambda} in tumour vascular networks following treatment with the anti-angiogenic cancer agent DC101. Based on our findings, we propose{lambda} as an effective way of monitoring the efficacy of antiangiogenic agents and as a proxy measure of perfusion and oxygenation in tumour tissue undergoing anti-angiogenic treatment. Significance statementOxygen heterogeneity in solid tumours is recognised as a limiting factor for therapeutic efficacy. This heterogeneity arises from the abnormal tumour vascular structure. We investigate the role that anomalies in RBC transport play in establishing oxygen heterogeneity in tumour tissue. We introduce a metric to characterise tumour vasculature (mean vessel length-to-diameter ratio,{lambda} ) and demonstrate how it predicts tissue oxygen heterogeneity. We also report an increase in{lambda} following treatment with the antiangiogenic agent DC101. Together, we propose{lambda} as an effective way of monitoring the action of anti-angiogenic agents and a proxy measure of oxygen heterogeneity in tumour tissue. Unravelling the causal relationship between tumour vascular structure and tissue oxygenation will pave the way for new personalised therapeutic approaches.

cancer biology↗

bioRxiv2019

Human neutrophils produce antifungal extracellular vesicles against Aspergillus fumigatus

Polymorphonuclear granulocytes (PMNs) are indispensable for controlling life-threatening fungal infections. In addition to various effector mechanisms, PMNs also produce extracellular vesicles (EVs). Their contribution to antifungal defense has remained unexplored. We reveal that the clinically important human pathogenic fungus Aspergillus fumigatus triggers PMNs to release a distinct set of antifungal EVs (afEVs). Proteome analyses indicated that afEVs are enriched in antimicrobial proteins. The cargo and release kinetics of EVs are modulated by the fungal strain confronted. Tracking of afEVs indicated that they associated with fungal cells and even entered fungal hyphae, resulting in alterations in the morphology of the fungal cell wall and dose-dependent antifungal effects. Two human proteins enriched in afEVs, cathepsin G and azurocidin, were heterologously expressed in fungal hyphae, which led to reduced fungal growth relative to a control retinol binding protein 7 producing strain. In conclusion, the production of afEVs by PMNs offers an intriguing, previously overlooked mechanism of antifungal defense against A. fumigatus. ImportanceInvasive fungal infections caused by the mold Aspergillus fumigatus are a growing concern in the clinic due to the increasing use of immunosuppressive therapies and increasing antifungal drug resistance. These infections result in high mortality as treatment and diagnostic options remain limited. In healthy individuals, neutrophilic granulocytes are critical for elimination of A. fumigatus from the host; however, the exact extracellular mechanism of neutrophil-mediated antifungal activity remains unresolved. Here, we present a mode of antifungal defense employed by human neutrophils against A. fumigatus not previously described. We find that extracellular vesicles produced by neutrophils in response to A. fumigatus infection are able to associate with the fungus, limit growth, and elicit cell damage by delivering antifungal cargo. In the end, antifungal extracellular vesicle biology provides a significant step forward in our understanding of A. fumigatus host pathogenesis and opens up novel diagnostic and therapeutic possibilities.

microbiology↗

bioRxiv2018

Tissue- and cellular-level allocation of autotrophic and heterotrophic nutrients in the coral symbiosis -- A NanoSIMS study

Corals access inorganic seawater nutrients through their autotrophic endosymbiotic dinoflagellates, but also capture planktonic prey through heterotrophic feeding. Correlating NanoSIMS and TEM imaging, we visualize and quantify the subcellular fate of autotrophic and heterotrophic C and N in the coral Stylophora pistillata using stable isotopes. Six scenarios were compared after 6h: autotrophic pulse (13C-bicarbonate, 15N-nitrate) in either unfed or regularly fed corals, and heterotrophic pulse (13C-, 15N-labelled brine shrimps) in regularly fed corals; each at ambient and elevated temperature. Host assimilation of photosynthates was similar under fed and unfed conditions, but symbionts assimilated 10% more C in fed corals. Photoautotrophic C was primarily channelled into host lipid bodies, whereas heterotrophic C and N were generally co-allocated to the tissue. Food-derived label was detected in some subcellular structures associated with the remobilisation of host lipid stores. While heterotrophic input generally exceeded autotrophic input, it was more negatively affected by elevated temperature. The reduced input from both feeding modes at elevated temperature was accompanied by a shift in the partitioning of C and N, benefiting epidermis and symbiont. This study provides a unique view on the nutrient partitioning in corals and highlights the tight connection of nutrient fluxes in symbiotic partners.

physiology↗

bioRxiv2018

A stochastic model of metastatic bottleneck predicts patient outcome and therapy response

Metastases are responsible for 90% of cancer-related deaths. Initiation of metastases, where newly seeded tumor cells expand into colonies, presents a tremendous bottleneck to metastasis formation. Despite its clinical importance, our understanding of this process is very limited. Here, we propose a simple stochastic model assuming that the initiating metastatic cells proliferate faster when surrounded by more of their kind. The model quantifies the severity of metastatic bottleneck as the probability that the seeded colony survives. Based on this model, we derive how metastasis occurrence depends on primary tumor size and affects patient outcome. Our predictions agree with epidemiological data for thirteen cancer types. The model predicts that impact of treatment decisions depends both on the primary tumor size and on the severity of the metastatic bottleneck, and that medical interventions that tighten the bottleneck would be much more efficient than therapies that decrease overall tumor burden, such as chemotherapy.

cancer biology↗

bioRxiv2017

Ancient symbiosis confers desiccation resistance to stored grain pest beetles

Microbial symbionts of insects provide a range of ecological traits to their hosts that are beneficial in the context of biotic interactions. However, little is known about insect symbiont-mediated adaptation to the abiotic environment, e.g. temperature and humidity. Here we report on an ancient (~400 Mya) clade of intracellular, bacteriome-located Bacteroidetes symbionts that are associated withgrain and wood pest beetles of the phylogenetically distant families Silvanidae and Bostrichidae. In the saw-toothed grain beetle Oryzaephilus surinamensis, we demonstrate that the symbionts affect cuticle thickness, melanization and hydrocarbon profile, enhancing desiccation resistance and thereby strongly improving fitness under dry conditions. Together with earlier observations on symbiont contributions to cuticle biosynthesis in weevils, our findings indicate that convergent acquisitions of bacterial mutualists represented key adaptations enabling diverse pest beetle groups to survive and proliferate under the low ambient humidities that characterize dry grain storage facilities.

evolutionary biology↗