Biology subjects

Gudelj, I.

Publications and source records attributed to Gudelj, I..

10 recordsLinked to original sources

bioRxiv2025

High-Throughput Site-Specific N-Glycosylation Profiling of Human Fibrinogen in Atrial Fibrillation

Fibrinogen is a major plasma glycoprotein involved in blood coagulation and inflammatory responses. Alterations in its glycosylation have been implicated in various pathological conditions, yet its site-specific N-glycosylation profile remains largely unexplored in a clinical context. Here, we present a high-throughput LC-MS workflow for site-specific analysis of fibrinogen N-glycosylation using a cost-effective ethanol precipitation enrichment method. The method demonstrated good intra- and inter-plate repeatability (CV: 5% and 12%, respectively) and was validated through the first assessment of intraindividual temporal stability in healthy individuals, revealing consistent glycosylation patterns within individuals. Application to 181 atrial fibrillation (AF) patients and 52 healthy controls identified three gamma chain glycoforms significantly associated with AF. Most notably, increased levels of the asialylated N4H5, known to enhance fibrin bundle thickness and promote clot formation, suggest a potential mechanism linking glycosylation changes to the prothrombotic state in AF. Furthermore, fibrinogen sialylation showed strong associations with cardiovascular risk factors, including triglycerides, BMI, and glucose levels. Longitudinal analysis of 108 AF patients six months post-catheter ablation showed stability in the AF-associated glycan profile. Our findings establish fibrinogen glycosylation as a potential biomarker for cardiovascular conditions and demonstrate the utility of site-specific glycosylation analysis for clinical applications.

molecular biology↗

bioRxiv2024

Exaptation and de novo mutations transcend cryptic variations as drivers of adaptation in yeast

Many organisms live in predictable environments with periodic variation in growth condition which can allow populations to accumulate cryptic genetic variations. Cryptic variations can facilitate adaptation to new environments, as observed in evolution experiments with a ribozyme and a protein. Whether the same holds for cell populations remains unclear. Alternatively, living in a near-constant condition can lead to loss of nonessential cellular functions, which could be maladaptive in new environments. Through laboratory evolution experiments in yeast, we show that populations grown in a predictable nutrient-rich environment for 1000 generations start to lose their ability to respond and adapt to new stressful environments. Growth of yeast populations in the nutrient-rich environment was associated with modest fitness increase in this environment, metabolic remodeling, and increased lipid accumulation. In novel stressful environments, however, these populations generally had reduced fitness, except in salt-stress where lipid accumulation seemed to provide osmotic protection. We further found that adaptation to stressors was primarily driven by de novo mutations, with very little contribution from the mutations accumulated prior to the exposure to stressors. Thus, our work suggests that in the absence of occurrence of new environments, natural populations might not accumulate cryptic variations that could be beneficial for adaptation to these environments. In addition, presence of selection in predictable condition in natural populations may purge away some of the cryptic variations. Taken together, these findings raise questions about persistence of cryptic variations in natural populations and their importance in evolutionary adaptation.

evolutionary biology↗

bioRxiv2023

METABOLIC MODELLING AS A POWERFUL TOOL TO IDENTIFY CRITICAL COMPONENTS OF THE PNEUMOCYSTIS GROWTH MEDIUM

Establishing suitable in vitro culture conditions for microorganisms is crucial for dissecting their biology and empowering potential applications. However, a significant number of bacterial and fungal species, including Pneumocystis jirovecii, remain unculturable, hampering research efforts. P. jirovecii is a deadly pathogen of humans that causes life-threatening pneumonia in immunocompromised individuals and transplant patients. Despite the major impact of Pneumocystis on human health, we remain ignorant about the pathobiology of this fungus. This is largely due to the fact that its experimental dissection has been constrained by the inability to culture the organism in vitro. We present a comprehensive in silico genome-scale metabolic model of Pneumocystis growth and metabolism, with a view to identifying metabolic requirements and imbalances that hinder growth in vitro. We utilise recently published genome data and available information in the literature as well as bioinformatics and software tools to develop and validate the model. In addition, we employ Flux Balance Analysis and Reinforcement Learning approaches to make predictions regarding metabolic fluxes and to identify critical components of the Pneumocystis growth medium. Our findings offer insights into the biology of Pneumocystis and provide a novel strategy to overcome the longstanding challenge of culturing this pathogen in vitro.

systems biology↗

bioRxiv2023

Human-specific features and developmental dynamics of the brain N-glycome

Comparative "omics" studies have revealed unique aspects of human neurobiology, yet an evolutionary perspective of the brain N-glycome is lacking. Here, we performed multi-regional characterization of rat, macaque, chimpanzee, and human brain N-glycomes using chromatography and mass spectrometry, then integrated these data with complementary glycotranscriptomic data. We found that in primates the brain N-glycome has evolved more rapidly than the underlying transcriptomic framework, providing a mechanism for generating additional diversity. We show that brain N-glycome evolution in hominids has been characterized by an increase in complexity and (2-6)-linked N-acetylneuraminic acid along with human-specific cell-type expression of key glycogenes. Finally, by comparing the prenatal and adult human brain N-glycome, we identify region-specific neurodevelopmental pathways that lead to distinct spatial N-glycosylation profiles in the mature brain. One-Sentence SummaryEvolution of the human brain N-glycome has been marked by an increase in complexity and a shift in sialic acid linkage.

neuroscience↗

bioRxiv2020

Clinical Antibiotic Resistance Patterns Across 70 Countries

We sought global patterns of antibiotic resistant pathogenic bacteria within the AMR Research Initiative database, Atlas. This consists of 6.5M clinical minimal inhibitory concentrations (MICs) observed in 70 countries in 633k patients between 2004 and 2017. Stratifying MICs according to pathogens (P), antibiotics (A) and countries (C), we found that the frequency of resistance was higher in Atlas than other publicly available databases. We determined global MIC distributions and, after showing they are coherent between years, we predicted MIC changes for 43 pathogens and 827 pathogen-antibiotic (PAs) pairings that exhibit significant resistance dynamics, including MIC increases and even decreases. However, many MIC distributions are multi-modal and some PA pairs exhibit sudden changes in MIC. We therefore analysed Atlas after replacing the clinical classification of pathogens into susceptible, intermediate and resistant with an information-optimal, cluster-based classifier to determine subpopulations with differential resistance that we denote S and R. Accordingly, S and R clusters for different PA pairs exhibit signatures of stabilising, directional and disruptive selection because their respective MICs can have different dynamics. Finally, we discuss clinical applications of a (R, dR/dt) phase plane whereby the MIC of R is regressed against change in MIC (dR/dt), a methodology we use to detect PA pairs at risk of developing clinical resistance.

microbiology↗

bioRxiv2020

Molecular Diversity Among Adult Hippocampal and Entorhinal Cells

The hippocampal-entorhinal system is comprised of functionally distinct subregions collectively critical for cognition, and selectively vulnerable to aging and pathological processes. To gain insights into neuronal and non-neuronal populations within this system, we performed single-nucleus transcriptional profiling from five human hippocampal-entorhinal subregions. We found that transcriptomic diversity of excitatory neurons across these subregions reflected the molecular transition from three-layered archicortex to six-layered neocortex. Additionally, mRNA and protein for DCX, an immature neuron marker, were clearly detected in some cells, but not in dentate granule cells, the cell-type predicted to be generated in adult neurogenesis. We also found that previously functionally uncharacterized METTL7B was enriched in human and non-human primate neuronal subtypes less vulnerable to initial Alzheimers disease pathology. Proteomic and biochemical assays revealed METTL7B interacts with Alzheimers disease-related proteins, including APP, and its overexpression reduced amyloid-beta generation. These results reveal cell type-specific molecular properties relevant for hippocampal-entorhinal physiology and dysfunction.

neuroscience↗

bioRxiv2019

Hotspot Dosages of Most Rapid Antibiotic Resistance Evolution

We treated Escherichia coli with the antibiotic erythromycin from zero to high dosages to determine how the evolutionary dynamics of antibiotic resistant phenotypes and genotypes depend on dose. The most rapid increase in resistance was observed just below erythromycins minimal inhibitory concentration (MIC) and genotype-phenotype correlations determined from whole genome sequencing revealed the molecular basis of this: simultaneous selection for copy number variation in 3 resistance mechanisms which shared an inverted-U pattern of dose-dependent selection with several insertion sequences and an integron. Many genes did not conform to this pattern, however, because of changes in selection as dose increased: media adaptation at zero-to-low dosages gave way to drug target (ribosomal RNA operon) amplification at mid dosages whereas prophage-mediated drug efflux dominated at higher dosages where population densities were lowest. All dosages saw E. coli amplify the efflux operons acr and emrE at rates that correlated strongly with changes in population density that exhibited an inverted-U geometry too. However, we show by example that inverted-U geometries are not a universal feature of dose-resistance relationships.

evolutionary biology↗

bioRxiv2019

Canonical host-pathogen tradeoffs subverted by mutations with dual benefits

Tradeoffs between life history traits impact diverse biological phenomena, including the maintenance of biodiversity. We sought to study two canonical tradeoffs in a model host-parasite system consisting of bacteriophage lambda and Escherichia coli: i) parasite resistance for growth and ii) phage infectivity for host-range. We report that these previously hypothesised tradeoffs are, in fact, tradeups. While the observation of tradeups was surprising, they should be expected because if traits X and Y tradeoff, so too traits Y and Z, then X and Z will tradeup. By considering five different E. coli trait correlations we uncovered several tradeups and tradeoffs. Using mathematical models, we establish that tradeups need not inhibit biodiversity, as previously thought, and can help maintain it through high-dimensional trait interactions. We provide a mechanistic explanation for how tradeups emerge and give reasons for why tradeups can even evolve in well-adapted genomes.\n\nAll data will be posted at https://github.com/rebear217 and mirrored at http://people.exeter.ac.uk/reb217/rebHomePage/data.html on acceptance.

evolutionary biology↗

bioRxiv2019

Fluorescence photography of patterns and waves of bacterial adaptation at high antibiotic doses

Fisher suggested advantageous genes would spread through populations as a wave so we sought genetic waves in evolving populations, as follows. By fusing a fluorescent marker to a drug efflux protein (AcrB) whose expression provides Escherichia coli with resistance to some antibiotics, we quantified the evolution and spread of drug-resistant E. coli through spacetime using image analysis and quantitative PCR. As is done in hospitals routinely, we exposed the bacterium to a gradient of antibiotic in a disk diffusion drug susceptibility test that we videoed. The videos show complex spatio-genomic patterns redolent of, yet more complex than, Fishers predictions whereby a decelerating wave front of advantageous genes colonises towards the antibiotic source, forming bullseye patterns en route and leaving a wave back of bacterial sub-populations expressing AcrB at decreasing levels away from the drug source. qPCR data show that E. coli sited at rapidly-adapting spatial hotspots gain 2 additional copies of acr, the operon that encodes AcrB, within 24h and imaging data show resistant sub-populations thrive most near the antibiotic source due to non-monotone relationships between inhibition due to antibiotic and distance from the source. In the spirit of Fisher, we provide an explicitly spatial nonlinear diffusion equation that exhibits these properties too. Finally, linear diffusion theory quantifies how the spatial extent of bacterial killing scales with increases in antibiotic dosage, predicting that microbes can survive chemotherapies that have been escalated to 250x the clinical dosage if the antibiotic is diffusion-limited.

evolutionary biology↗

bioRxiv2019

Global variability of the human IgG glycome

Immunoglobulin G (IgG) is the most abundant serum antibody and is a key determinant of the humoral immune response. Its structural characteristics and effector functions are modulated through the attachment of various sugar moieties called glycans. IgG N-glycome patterns change with the age of individual and in different diseases. Variability of IgG glycosylation within a population is well studied and is affected by a combination of genetic and environmental factors. However, global inter-population differences in IgG glycosylation have never been properly addressed. Here we present population-specific N-glycosylation patterns of whole IgG, analysed in 5 different populations totalling 10,482 IgG glycomes, and of IgGs fragment crystallisable region (Fc), analysed in 2,530 samples from 27 populations sampled across the world. We observed that country of residence associates with many N-glycan features and is a strong predictor of monogalactosylation variability. IgG galactosylation also strongly correlated with the development level of a country, defined by United Nations health and socioeconomic development indicators. We found that subjects from developing countries had low IgG galactosylation levels, characteristic for inflammation and ageing. Our results suggest that citizens of developing countries may be exposed to country-specific environmental factors that can cause low-grade chronic inflammation and the apparent increase in biological age.

biochemistry↗