PhD Projects
Call for applications from Doctoral Candidates - NOW OPEN!
For more details and to apply, follow the links with each individual project below.
The GLYCOCALYX Doctoral Network will provide 15 doctoral candidates with training in bespoke physics, chemistry and biology methods – essential disciplines that will be integrated to enable us to resolve the dynamic organisation of glycocalyces, and how they perform the many selective barrier functions essential to multicellular life.
DC1: Biocolloid diffusion and transport in glycocalyces
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Host Institution: University of Leeds (UK)
Main Academic Supervisor: Ralf Richter
Click HERE (Link coming soon) to apply for DC1 position
The glycocalyx ‘forest’ accomplishes many crucial roles in the communication of cells with their environment by controlling cell-surface access of a wide range of ‘biocolloids’, including signalling proteins (e.g., chemokines, growth factors, morphogens), extracellular vesicles, and pathogenic agents (toxins, viruses). Whilst the ultimate biological effect of each biocolloid is different, there are common molecular features that control the transport across the glycocalyx: biocolloid size relative to glycocalyx pore size sterically control permeation, and (often multiple) weak biocolloid-glycocalyx interactions modulate enrichment and transport rates. The universal physics rules that define how this combination of features regulates biocolloid partitioning and diffusion, however, are not well understood.
This project will combine expertise in molecularly defined model glycocalyces and advanced microscopy methods to probe biocolloid diffusion and enrichment in model glycocalyces with defined and tuneable pore size and multivalent interaction characteristics. The project will define universal physics rules of biocolloid diffusion and enrichment by confronting experimental data with theory and computer simulations (in collaboration with Network partners). These rules will enable the rational design of regulators of glycocalyx permeation, to interfere with pathogen-host cell interactions and for targeted delivery into cells.​
DC2: Glycocalyx self-organisation: the driving force for perineuronal net formation?
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Host Institution: University of Leeds (UK)
Main Academic Supervisor: Jessica Kwok
Click HERE to apply for DC2 position
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Perineuronal nets (PNNs) are specialised, hierarchical assemblies of the extracellular matrix that surround subsets of neurons and play a critical role in regulating neuronal plasticity in the adult central nervous system. In the mature brain, PNNs form distinctive reticular structures, with openings that accommodate synaptic contacts. Modulation of PNNs has been shown to influence plasticity and functional recovery in a range of neurological contexts, highlighting their importance for learning, memory, and brain repair. Despite their functional significance, the mechanisms by which PNN components organise into the characteristic PNN architectures remain poorly understood. This PhD project aims to address this knowledge gap by investigating how interactions between PNN molecules give rise to their ultrastructural organisation.
The project will adopt an interdisciplinary experimental approach to explore how PNN composition and molecular interactions influence their spatial organisation and structural properties on the neuronal surface. You will apply advanced super-resolution microscopy, neuronal culture, protein expression and labelling, molecular interaction assays, in conjunction with physics-based analysis and modelling, to determine how PNNs form. By integrating insights from extracellular matrix biology, glycobiology, neurobiology and biophysics, the research will contribute new insights into PNN organisation and function, with long‑term relevance for strategies to modulate neuroplasticity.
DC3: Targeting tissues using multivalent lectin-glycan interactions
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Host Institution: University of Leeds (UK)
Main Academic Supervisor: Bruce Turnbull
Click HERE (Link coming soon) to apply for DC3 position
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Many cell types express a similar set of glycans, and it is the relative abundance of glycan epitopes, rather than the mere presence of a given epitope, that differentiates one cell type from another. This project aims to develop ‘superselective’ glycocalyx probes that can recognise specific glycan epitopes in a density-dependent fashion. The resulting probes will have applications in super-resolution microscopy of glycocalyx organisation, and for targeted delivery of molecules with exquisite selectivity. We have demonstrated that multivalent probes based on a flexible polymer scaffold can effectively discriminate surfaces based on the density of surface receptors. In this project you will develop new polymer scaffolds, and use enzymatic and bio‑orthogonal chemistry to precisely functionalise lectins to assemble the modular probes. Their binding behaviour will be studied in model glycocalyces using QCM‑D, and their interactions with neuronal and endothelial cells will be visualised by super‑resolution microscopy. The work combines chemical biology, glycoscience, theoretical modelling, and advanced imaging in an interdisciplinary research environment.
DC4: Imaging the dynamics and viscoelastic properties of the glycocalyx at single molecule resolution
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Host Institution: TU Wien (Austria)
Main Academic Supervisor: Gerhard Schütz
Click HERE to apply for DC4 position
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Microrheology is an emerging tool to study viscoelastic properties of the glycocalyx on live cells. By tracking single particles of different size and analysing diffusional motion, it is possible to infer the elastic modulus and viscosity, as well as the apparent thickness, of the glycocalyx. Currently, however, this method is largely restricted to the interface between adherent cells and glass coverslips, which is likely affected by the rigid glass counter-surface. Specific aims are: 1) Develop TIRF-based 2-dimensional single particle tracking microrheology to determine frequency-rate-dependent viscoelastic moduli in the contact area of dendritic cells with glass coverslips; 2) Use single molecule sensitive (lattice) light sheet microscopy in three dimensions to investigate glycocalyx viscoelasticity on the dorsal surface of dendritic cells and T-cells; 3) Access glycocalyx dynamics at the nanometre scale by tracking the motion of fluorescently labelled glycocalyx motifs; 4) Probe glycocalyx dynamics in the immunological synapse between T-cells and antigen-presenting dendritic cells. The project will benefit from single particle tracking and other dynamic optical microscopy, glycocalyx cell models, and glycocalyx labelling tools in the network.
DC5: Impact of selective shedding of the endothelial glycocalyx on haemodynamics
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Host Institution: Laboratory of Interdisciplinary Physics, CNRS, Grenoble (France)
Main Academic Supervisor: Lionel Bureau
Click HERE to apply for DC5 position
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The endothelial glycocalyx (EG), which lines the lumen of blood vessels, is a key player in regulating interactions between circulating blood cells and the vessel walls. It is a micron-thick hydrated meshwork comprising glycosaminoglycans (GAGs), such as heparan sulfate (HS), chondroitin sulfate (CS) and hyaluronan (HA), as its main constituents. The loss of structural integrity and shedding of EG components are recognised to be associated with multiple acute or chronic vascular pathologies, and to affect hemodynamic features such as repulsion of red blood cells by the vessel walls or adhesion of immune cells. However, the respective role of HS, HA and CS in controlling near-wall haemodynamics is not well understood, and whether they act synergistically to control EG properties and functions is essentially unknown. We will address these questions by performing in vitro experimental studies using endothelialised microfluidic devices developed in our lab. ​Specific aims are: 1) Assess the GAG composition of the glycocalyx expressed by endothelial cells cultured under physiologically relevant shear flow; 2) Evaluate the impact of selective enzymatic degradation of the three main GAGs on glycocalyx thickness; 3) Study how such selective shedding affects elastohydrodynamic and adhesive interactions between the endothelium and circulating blood cells; 4) Assess how the mechanical properties of the EG control such interactions. The project will combine glycocalyx cell models, dynamic optical microscopy, glycocalyx labelling and GAG enzyme biochemistry.​
DC6: Glycocalyx softness and control of bacterial adhesion
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Host Institution: Laboratory of Interdisciplinary Physics, CNRS/UGA, Grenoble (France)
Main Academic Supervisor: Delphine Débarre
Click HERE to apply for DC6 position
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Pathogenic bacteria such as P. aeruginosa sense their biochemical and mechanical environments. We and others have recently demonstrated that surface colonisation by such bacteria is strongly affected by surface elasticities in the kPa range. Conversely, mammalian glycocalyces act as gate keepers controlling pathogen colonisation of endothelia and epithelia, and their extreme softness (1-100 Pa range) may play a key role in regulating bacterial adhesion. How bacterial cells integrate mechanical cues on such soft, fuzzy interfaces is, however, unknown. Using our combined expertise in building well-defined model glycocalyces in microfluidic devices, and in monitoring bacterial adhesion and biofilm formation, we will study how the softness of model glycocalyces impacts the early stages of surface colonisation and the onset of biofilm formation. The project will combine molecularly defined glycocalyx models and microfluidics, dynamic optical microscopy, bacterial biophysics, and theoretical modelling of soft matter systems.
We will define the key glycocalyx physical properties that regulate bacterial colonisation. This will advance our understanding of the role of a healthy and diseased glycocalyx in the early stages of bacterial infection, and provide new avenues for the design of biomimetic anti-fouling surfaces.​​​
DC7: Recognition and self-organisation of the glycocalyx – Does binder clustering matter?
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Host Institution: Department of Molecular Chemistry, CNRS/UGA, Grenoble (France)
Main Academic Supervisor: Galina Dubacheva
Click HERE to apply for DC7 position
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Transient multivalent interactions are commonplace in glycocalyces. They control the recognition between the glycocalyx and signalling proteins, extracellular vesicles and pathogens (toxins, viruses), and also contribute to the supramolecular organisation of the glycocalyx itself (e.g., through macromolecular cross-linking). Multivalent interactions are inherently sensitive to the spatial disposition of binding sites, but how the distribution of binders affects glycocalyx recognition and self-organisation is not well understood. The complexity and poor control over the composition and organisation of cellular glycocalyces makes such studies challenging. The goal of this project is to elucidate how clustering of binders, as opposed to a homogeneous binder distribution, impacts glycocalyx recognition and physical properties. ​Specific aims are: 1) Develop chemical tools to make synthetic ‘designer’ glycocalyces with defined and tuneable binder affinities, concentrations and distributions (clustered vs. homogeneous); 2) Use synthetic glycocalyces to probe how binder clustering affects multivalent binding to a soft and fuzzy interface; 3) Use synthetic glycocalyces to probe the effect of cross-linker valency on the physical characteristics of glycocalyx-like films (e.g., phase separation, permeability). The project will combine bioorganic synthesis and host/guest supramolecular chemistry, surface functionalisation and physicochemical characterisation, theoretical/computational analyses and validation on biological systems through the network.
DC8: Spatial fingerprinting of nanoscale glycocalyx architectures
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Host Institution: Friedrich Alexander University and University Hospital Erlangen (Germany)
Main Academic Supervisor: Leonhard Möckl
Click HERE (Link coming soon) to apply for DC8 position
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Research insights established over the past years suggest that glycocalyx state has a direct connection to cell function, both in health and disease. For example, immune system regulation is tightly linked to the interactions between sialylated species and Siglecs, mostly inhibitory membrane receptors found on immune cells. Furthermore, various cancer types present aberrant glycosylation patterns on the cell surfaces, for example overexpression of large, bulky species like MUC1 or MUC16. Such pathological changes are suspected to aid cancer progression and metastasis. However, the mechanisms by which these physiological and pathological glycosylation phenotypes regulate cell function are only poorly understood. A key reason for this is that we lack information about the spatial presentation and organisation of these glycans in the cellular environment. In particular, the structural complexity and small spatial dimensions of the glycocalyx make access to such information challenging. The Möckl lab has recently established methods based on metabolic labelling, bio-orthogonal chemistry and super-resolution microscopy for the visualisation of many characteristic glycan structures with nanometer resolution on cells. Specific aims are: 1) Expand and refine current protocols towards highly multiplexed, molecular-resolution microscopy of glycans; 2) Apply them to precisely defined cellular model systems of relevant cancer types, such as conditionally activatable proto-oncogenes and models of the epithelial-to-mesenchymal transition; 3) Establish an atlas of cancer-related glycosylation patterns at the nanoscale. The project will combine expertise in super-resolution microscopy and glycocalyx labelling tools.​
DC9: Lectin-based labels for glycan imaging using DNA-PAINT super-resolution microscopy
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Host Institution: Massive Photonics GmbH, Munich (Germany)
Main Academic Supervisor: Sebastian Strauss
Click HERE (Link coming soon) to apply for DC9 position
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The ability to visualise glycans with high specificity and resolution is crucial for understanding their role in various biological processes and diseases. Current glycocalyx analysis is restricted by the lack of probes that specifically and efficiently target glycans. The goal of this project is to develop lectin-based labels for specific and quantitative glycan imaging.​ Specific aims are: 1) Establish conjugates of commercially sourced lectins with fluorescent dye (for conventional fluorescence microscopy) and DNA (for DNA-PAINT microscopy), and screen for their suitability for cellular glycocalyx imaging; 2) Selecting the best hits, recombinantly express new lectin variants with site-specific labelling tags for quantitative super-resolved glycan visualisation. Lectin conjugates will be screened for their glycan binding kinetics by BLI or SPR, followed by two-step imaging validation: conventional diffraction-limited microscopy with lectin-dye conjugates followed by super-resolution DNA-PAINT microscopy using lectin-DNA conjugates.
DC10: The mucosal glycocalyx and the host-microorganism relationship in homeostasis and infection
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Host Institution: i3S, Porto (Portugal)
Main Academic Supervisor: Salomé Pinho
Click HERE to apply for DC10 position
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Glycans are essential components of our immunological identity, fundamental for the discrimination between self and non-self, which relies on glycan recognition by glycan-binding proteins (lectins, antibodies) expressed or secreted by immune cells. The importance of the mucosal glycocalyx in the relationship between microorganisms – commensal or pathogenic – and their host, and most notably in the regulation of the host immune system, has only recently become appreciated. We are currently unable to decipher how the vast information displayed by the glycocalyx, and its spatiotemporal regulation in health and in infection/inflammation, define the magnitude, the nature and the fate of immune responses both in homeostasis and in disease. Specific aims are: 1) Investigate how mucosal glycocalyx composition and spatial organisation define the mucosal microenvironment, i.e., interactions with the microbiome and the immune system; 2) Investigate how glycan recognition by selected glycan-binding host proteins impacts homeostasis and immunological tolerance; 3) Investigate how changes in mucosal glycocalyx composition/organisation define susceptibility to infection (by selected bacteria, viruses and/or fungi) leading to a modulation of the immune response and disease onset. The project will benefit from expertise in glycoimmunology with access to unique glycoengineered in-vitro, ex-vivo and in-vivo models and human samples, glycomics, microscopy and glycocalyx labelling tools.
DC11: Synthetic dendritic cell glycocalyx – Bridging the gap between molecules and cells
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Host Institution: CIC biomaGUNE, San Sebastian (Spain)
Main Academic Supervisor: Natalia Baranova
Click HERE to apply for DC11 position
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Even though the glycocalyx coat can be found on nearly every living cell across kingdoms of life, the relationship between its architecture, dynamic re-organisation and cell function remains largely unexplored. Dendritic cells (DCs) are best known for their role as professional antigen-presenting cells in T-cell antigen recognition. This process is characterised by a remarkable sensitivity combined with enormous selectivity: T-cells can specifically detect via their T-cell antigen receptor (TCR) the presence of a few down even to single antigenic peptides presented via MHC at the surface of an antigen-presenting cell, even at a background of thousands of structurally similar yet non-activating peptides. During the search for antigen, T-cells first tiptoe the surface of DCs with their microvilli extensions. To reach the plasma membrane, microvilli have to penetrate the glycocalyx coat. In this project, in a tight collaboration with Gerhard Schütz group (TU Wien), we will reconstitute components of the DCs glycocalyx on a fluid lipid membranes and investigate their effect on the T-cell interaction with the surface, microvilli dynamics and signaling activation. Direct correlation between cellular and reconstituted assemblies will enable us to gain a mechanistic understanding of the underlying molecular processes that are otherwise masked by cellular complexity and fast dynamics of the interactions. The reverse engineering of DCs glycocalyx will provide important information for immune cell reprogramming. Considering the accumulation of the extensive glycocalyx around cancer cells, the synthetic glycocalyx can also be used as a platform to improve the design of synthetic chimeric antigen receptors (CARs) T-cells.
DC12: Viral infections in the endothelium – Glycocalyx penetration and viral pathogenesis
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Host Institution: Umeå University (Sweden)
Main Academic Supervisor: Marta Bally
Click HERE to apply for DC12 position
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To infect a host cell, viruses need to be transported through the glycocalyx to reach the cell membrane. This is particularly challenging for viruses known to infect directly the endothelium, since endothelial cells carry a particularly thick (up to μm) glycocalyx. On a molecular level, it remains poorly understood how these viruses have evolved to breach this barrier, while interacting with its molecular components. It is also unclear whether there is a relationship between the efficiency with which viruses cross the glycocalyx and their ability to cause disease. In this project, we will work with viruses infecting the endothelium. Specific aims are: 1) Investigate how diffusion potential within the glycocalyx relates to its molecular features (e.g., binding site densities and molecular architecture); 2) Define whether different viruses of the same virus family but with distinct pathogenic potential differ in their ability to cross the endothelial glycocalyx. This project will deploy molecularly defined model glycocalyces, in vitro cell models with a thick glycocalyx, single particle tracking and (live cell) microscopy.
DC13: Endothelial glycocalyx degradation during viral diseases
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Host Institution: Umeå University (Sweden)
Main Academic Supervisor: Marta Bally and Anne-Marie Fors Connolly
Click HERE to apply for DC13 position
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Endothelial glycocalyx (EG) degradation has been associated with a multitude of vascular pathologies, including cardiovascular complications associated with viral diseases. This process is mediated by ‘sheddase’ enzymes which degrade EG components thereby affecting EG biophysical properties (e.g., thickness, stiffness and permeability). Recently, two viral diseases, potentially leading to severe cardiovascular complications, have been shown to associate with EG degradation: haemorrhagic fever with renal syndrome (HFRS) caused by Puumulavirus (PUUV; a hantavirus), and COVID-19 caused by SARS-CoV-2. In both cases, it remains unclear whether there is a direct correlation between EG degradation, pathogenesis and disease severity. The molecular mechanisms at play also remain to be elucidated. Specific aims are: 1) Gain fundamental understanding of the molecular mechanism underlying EG degradation during PUUV and SARS-CoV-2 infection; 2) Establish a correlation between disease stage/severity and EG biophysical properties. To this end, we will deploy composition and physical property analyses with an established in vitro cell model. Studies will be carried out using relevant purified sheddases, and blood plasma samples from HFRS and COVID-19 patients stratified across disease stages and severity, in the presence or absence of sheddase inhibitors.​
DC14: Analysis of glycocalyx glycans using novel LC-FD-MS chromatography techniques
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Host Institution: Ludger Ltd. (UK)
Main Academic Supervisor: Daniel Spencer
Click HERE (Link coming soon) to apply for DC14 position
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The mammalian glycocalyx displays N- and O-linked glycans with a wide range of complexity, branching, and isomerisation patterns. A standard method for analysing these glycans, with excellent repeatability, is hydrophilic interaction liquid chromatography (HILIC), which detects released and fluorophore-derivatised (FD) glycans. In-depth studies often employ exoglycosidase reactions to resolve some of the complexity, but these are impractical for large-scale analyses involving multiple samples. Previously, we used HILIC-FD coupled to mass spectrometry (MS) to analyse cell surface glycosylation, revealing multiple isomers, along with extensive sialylation and fucosylation. Building on this, we have now developed a C18-based liquid chromatography (LC)-FD-MS method combining procainamide (proc) fluorescence (for relative quantitation of glycans and efficient ionisation for MS detection) with ethyl esterification (EE; for discrimination of immunity/infection relevant 2-3 and 2-6 sialic acid linkages), thus enabling the identification of glycan isomers previously hard to define. C18 chromatography offers a wide range of column types, from preparative to nanoflow, including core-shell designs that support the high flow rates necessary for more rapid sample turnaround time. The goal of this project is to further refine the Proc/EE/C18 method to enable detailed characterisation of complex glycans, including those with isomerisation and diverse bond types. Specific aims are: 1) Enhance the Proc/EE/C18 method for more detailed glycan analysis focusing on column resolution and sensitivity; 2) Develop rapid LC techniques for higher-throughput sample processing (<20 min run times); 3) Apply these advancements to key immunology and infection research projects in collaboration with the network (e.g., Salomé Pinho, i3S).​​
DC15: Combining coarse-grained modelling and machine learning to unravel the role of the
glycocalyx in modulating viral attachment to the cell-surface
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Host Institution: Imperial College London (UK)
Main Academic Supervisor: Stefano Angioletti-Uberti
Click HERE (Link coming soon) to apply for DC15 position
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The glycocalyx acts as a selective barrier that viruses and other biological entities like bacteria or even proteins need to pass to reach the cell surface. In terms of physical interactions, a repulsive term arises from the need to penetrate through the dense glycocalyx polymer ‘brush’, while attraction is dictated by the various transient, reversible bonds formed by (viral) receptors and their cognate ligands, the latter constituted by specific glycans in the glycocalyx (e.g., sialic acids for influenza virus). This physical picture qualitatively illustrates that the free-energy profile that a ‘biocolloid’ (virus, protein or bacteria) encounters on its approach to the cell surface will depend on the structure of the glycocalyx and the corresponding multivalent presentation of ligands. Knowledge of this free-energy profile can be used to quantitatively link the chemistry and structure of the glycocalyx to the kinetics of viral attachment to the cell, and thus to infectivity. We will provide such a link by combining concepts from polymer physics, the theory of multivalent interactions, kinetic theory and machine learning techniques. In parallel, we will also use deep-learning based tools to design molecular probes to characterise and tune glycocalyx/biocolloid interactions. Specific aims are: 1) Build a coarse-grained model of the glycocalyx/virus interaction, and study this model to build a general understanding of the underlying physics governing viral transport and attachment kinetics; 2) Use a combination of Monte Carlo simulations and a Machine Learning approach based on Bayesian optimisation via Gaussian Processes to explore the parameter space of the glycocalyx-virus system parameters, with the goal to find the regions maximising and minimising viral attachment kinetics; 3) Use deep-learning based tools to design molecular probes to characterise and tune glycocalyx/biocolloid interactions, and 3) Validate the computational model results by comparison to experimental data on model systems. The knowledge gained will provide rational principles and tools to understand the link between glycocalyx properties and viral infectivity, and also rational rules to design synthetic systems like nanoparticles for targeted drug delivery to cells.
This project has received funding from the European Union’s Horizon Europe research and innovation programme under grant agreement GLYCOCALYX – 101227305