The addition of sugars to a protein structure can affect protein stability, receptor interactions with ligands, and signaling pathway activity. This modification, called glycosylation, is linked to inflammation and disease progression. A new paper examines the understanding of glycosylation in a range of rheumatic diseases and indicates the potential for therapeutic interventions with the same mechanism.
Study: Glycobiology of rheumatic diseases. Image credit: zentradyi3ell/Shutterstock
Introduction
Small and large chains of sugar molecules, called oligo and polysaccharides, respectively, or together called glycans, are often added to fat or protein molecules to modify them. Half of human genes encode glycosylated proteins. Glycans are built by sequentially adding sugars through the coordinated activity of many enzymes. The final signature is made up of many sugars that have related structures but have quite different functions.
Glycans affect a number of cellular pathways, including communication between cells and the processes that take place in cells. The outer surface of cells is often covered by a dense glycan layer called the glycocalyx that is key to, but also influences, intercellular communications.
The current article, published in Nature Reviews Rheumatology, summarizes current knowledge on the glycosylation patterns of plasma proteins, autoantibodies, immune cells and inflamed tissues in rheumatic diseases, along with some mechanistic insights and suggestions for the therapeutic potential of these findings.
Both O- and N-glycosylation have been identified and occur through different pathways. The final signature is affected by the availability of sugars, enzymes, and transporters required for glycosylation and the extent to which the appropriate molecular regions are open to these reactions. Cellular state also affects glycan composition by modulating the entry or exit of glycoproteins into or out of the endoplasmic reticulum (ER) and the Golgi apparatus, both of which are involved in post-translational protein modification.
N-glycan changes
Glycans are involved in a wide range of immunological reactions, leading to the postulation that they are important in regulating the immune response during autoimmune rheumatic disease. For example, rheumatoid arthritis (RA) is associated with changes in the glycosylation pattern of immunoglobulin G (IgG) autoantibodies.
Serum proteins also show changes in glycosylation patterns in RA patients consistent with acute inflammatory changes and immune cell recruitment. These changes may also be related to disease activity, particularly with sialyl and fucosyl residues, while reduced activity is associated with galactosyl residues.
This is thought to be associated with changes in levels of IgG subclass autoantibodies directed against specific autoantigens in rheumatic diseases, including RA, psoriatic arthritis and many other arthropathies, but also multiple sclerosis and inflammatory bowel disease . These show a distinct profile of N-glycosyl residues in the antibody region called the crystallizable fragment (Fc) domain, marked by the presence of fucosyl but not sialyl or galactosyl residues.
Active RA shows a distinct pattern of glycan residues in both the Fc and Fab domains of IgG autoantibodies, while patients in remission show another specific profile. Absence or deletion of galactosyl residues in the Fc domain is associated with active, progressive or recurrent disease and could be a useful biomarker for rheumatic diseases. Galactosylation of IgG antibodies increases when RA patients become pregnant, indicating a phase of remission for many of these patients, but this is rapidly reversed after delivery.
However, an increase in sialyl and galactosyl residues is associated with reduced disease severity in RA, although this may not be specific to the Fc domain. Thus, “Total IgG [and] Antigen-specific IgG has Fc N-glycosylation that correlates with disease state.”
Both rheumatoid arthritis (RA) and anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis often test positive for another type of N-glycosylated autoantibody, as well as B-cell receptors that react with autoantigens . These contain many N-glycosyl residues in the antibody binding fragment (Fab) domain, which could accompany an immunomodulatory function. N-linked glycosylation of the Fab domain could be in response to chronic exposure to whole-body antigens, together with CD4 T cell activity facilitating the survival of autoreactive B cells.
These Fab glycosylation changes could facilitate autoimmune phenomena by helping autoreactive B cells to overcome immune tolerance.
Mechanisms of action
Future research should explore the mechanisms of inflammation associated with the presence of IgG Fc sugars that include fucosyl but not galactosyl residues. Some possibilities include steric hindrance by fucosyl residues in the Fc domain that prevents activation of the corresponding Fcγ receptor, FcγRIIIa, reducing binding affinity and cytotoxicity up to 20-fold. The same relationship was observed in coronavirus disease 2019 (COVID-19), underscoring the increased inflammation evoked by IgG antibodies lacking fucosyl residues against the spike protein of the causative virus.
In humans, a large number of galactosyl residues enhance binding to the initiator of the classical complement pathway, independent of fucosylation levels. This could be due to increased hexamer formation induced by this change in the Fc domain.
However, “no clear mechanistic evidence is yet available to confirm that agalactosylated Fc glycans exacerbate antibody effector functions in humans.” Other explanations are possible, including a glycation response to inflammation.
Glycosylation of tissues and cells
The glycocalyx is important for modifying cell membrane shape and receptor distribution, affecting signaling function, cell adhesion and migration, ultimately modifying immunotolerance in autoimmune diseases.
In RA, synovial fibroblasts show a poor concentration of surface sialyl residues in response to the altered cytokine pattern. This inhibits its binding to galectin-3, causing the release of a distinct pattern of cytokines characteristic of a pro-inflammatory switch.
After tumor necrosis factor (TNF) treatment, sialylation is reduced, consistent with disease remission. Conversely, TNF stimulation is associated with decreased sialylation and increased disease activity in RA patients.
“Together, these data provide the first evidence that, in rheumatic disease, sialylation can act as a ‘molecular switch’, controlling the inflammatory or resting state of synovial fibroblasts.”
Clinical implications
Identification of specific IgG glycan signatures could help improve prognostic accuracy, but this type of analysis is expensive and difficult. Further development of appropriate assays is therefore awaited.
Therefore, specific modification of the glycosylation profiles of proteins, either antibody or tissue, could be useful to treat the underlying inflammation, either by altering the glycosylation pattern or by inducing the specific binding and destruction of autoantibodies and cells B autoreactive. Cytokines could modify the activity of enzymes involved in glycosylation.
Again, specific glycans, enzyme substrates, or metabolites could be manipulated to alter glycosylation patterns. For example, N-acetyl glucosamine reduces chronic inflammation and autoimmune activity by increasing the hexosamine pathway. This supplement could prevent T-cell receptor hyperactivation by increasing the degree of N-glycan branching.
Glycosylation is also key in the new area of glycoengineered monoclonal antibodies developed for various biological applications. These modifications can optimize their effectiveness, for example, by improving their binding affinity.