Glycans as mediators of infection and inflammation

The majority of all proteins are modified with glycans. Glycans create structural and functional diversity for otherwise identical proteins, being expressed intracellularly, on cell surfaces or in the extracellular space. Glycan modifications are necessary since they interact specifically with endogenous lectins for both conformational changes, turn-over as well as signalling within and between cells. When the biosynthesis or degradation of these glycans goes wrong, for genetic or acquired reasons, dysfunctional and dysmorphic characteristics will appear either for the whole individual or for single cell types e.g. defective or suppressed immune response against infection and cancer. We specifically study how pathogenic viruses utilize host cell glycans to infect cells, spread and avoid the immune response.

Glycan modified proteins, i.e. glycoproteins, are often classified according to which type of glycans they are carrying and how these are linked to the core proteins. Traditionally, they are named either N -glycoproteins, since their glycans are linked to an asparagine residue (Asn; N) located in a specific amino acid sequence (N-X-T/S) or O-glycoproteins since their glycans are linked to either of the amino acids serine (Ser;S), threonine (Thr;T) or tyrosine (Tyr; Y). For the latter three there is no definite amino acid consensus sequence and, in comparison with the N-linked glycans, the O-glycans vary considerably more in their structures with everything from single monosaccharide residues to several hundreds placed aside each other (as in mucins) or along a long polysaccharide chain (as in proteoglycans). The strict differences between these various types of glycoproteins are partly being abandoned due to advances in the analytical techniques revealing many different kinds of glycan modifications on the same protein and also ever more glycan modifications on earlier defined non modified proteins (and peptides).

The biosynthesis of the complex glycans is carried out by a series of more than a hundred different enzymes and transport proteins, which act together in a manner not fully understood, mostly in the Golgi apparatus but also in the endoplasmic reticulum and in the cell cytoplasm. These enzymes are typically classified as initiating, extending and modifying and finally as capping enzymes and while the two former are unique to each type of glycan modification, the latter are often common to glycans found both on N- and O-glycoproteins and even on glycosphingolipids.

Our own research has for a long time being focussed on studying the interactions between the cell surface glycoconjugates and pathogenic bacteria and more lately enteric viruses. We were the first to develop in vitro techniques for identifying the specificities of various microorganisms binding to complex glycans of reference glycolipids, that we ourselves had prepared and structurally characterized in detail by mass spectrometry and NMR spectroscopy. More lately we have, much thanks to our tight collaboration with Fredrik Höök and his group at Chalmers University of Technology, advanced these binding studies to include QCMD and TIRF-M techniques to study the dynamics (affinity, avidity and diffusion) of non-enveloped virus binding to glycolipids in solid supported lipid membranes. In this way we have characterized in detail the binding of norovirus to intestinal glycosphingolipids and their biosynthetic dependence on a functional FUT2 gene, necessary for becoming susceptible to the globally dominating norovirus strains causing the winter vomiting disease. We now stand ready to study in real time the dynamics of virus binding, uptake and propagation in cultured cells.

In parallel to these studies we have been developing novel mass spectrometric methods for the structural characterization of all glycoproteins, most lately focusing on proteoglycans mapping both the core proteins, their glycosaminoglycan chains and their glycan attachment sites.  During this technological development we have identified several novel proteoglycans in both human, mouse, rat, zebrafish, Drosophila and C. elegans. In many of these systems, including humans, we have been able to show how differences in glycosylation directly alter the functional characteristics of the core proteins and leads to defects in cell and tissue differentiation.

We now intend to unite all our areas of expertise to study how enveloped viruses, such as Herpesviruses, Influenzaviruses and Coronaviruses, interact with glycoproteins on the cell surface, how the viruses are taken up by the cells, how their genomes are released into the cytoplasm in the process of “endosomal escape”, are translated into viral proteins and glycoproteins and finally the egress and spread of mature viruses. These viruses have different specificities for host cell glycans but specific focus will be given to SARS-CoV-2, which binds to the ACE receptor, but which also needs an interaction with heparan sulfate proteoglycans to bind stronger to the cell surface and to more efficiently be taken up by the cells. The surface glycoproteins of enveloped viruses are often covered with host glycans as a ”memory” of the cells in which they have been produced and these modifications serve as glycan shields avoiding the host immune response, i.e. the immune cells do not recognize the virus surface as foreign. It is our goal to understand at the molecular level, the mechanisms behind these intricate interactions between the viruses and the host cells (infected cells and immune cells) in order to develop inhibitory low molecular weight compounds, antibodies and efficient vaccines activating the cellular immune response and defeating these pathogens.