Citrullination only infrequently impacts peptide binding to HLA class II MHC

Abstract

It has been hypothesized that HLA class II alleles associated with rheumatoid arthritis (RA) preferentially present self-antigens altered by post-translational modification, such as citrullination. To understand the role of citrullination we tested four RA-associated citrullinated epitopes and their corresponding wild-type version for binding to 28 common HLA class II. Binding patterns were variable, and no consistent impact of citrullination was identified. Indeed, in one case citrullination significantly increased binding compared to the WT peptide, in another citrullination was associated with a reduction in promiscuity by 40%. For a more comprehensive analysis, we tested over 200 citrullinated peptides derived from vimentin and collagen II for their capacity to bind the RA-associated shared epitope alleles DRB1*01:01 and DRB1*04:01. The overall effect of citrullination on binding was found to be relatively minor, and only rarely associated with 3-fold increases or decreases in affinity. Previous studies have suggested that citrullination of MHC anchor residues, in particular P4, is associated with generation of novel RA-associated epitopes. However, analysis of the predicted MHC-binding cores of all peptides tested found that in modified peptides with increased binding affinity the citrullinated residue was predicted to occupy an anchor position in only a minority of cases. Finally, we also show that identification of citrullinated peptide binders could be facilitated by using the NetMHCIIpan 3.1 algorithm, representing citrullination as a wildcard. Our studies identify a total of 117 citrullinated peptides that bound RA-associated alleles with an affinity of 1000 nM or better.

T cell repertoire to citrullinated self-peptides in healthy humans is not confined to the HLA-DR SE alleles; Targeting of citrullinated self-peptides presented by HLA-DP4 for tumour therapy

“…19,20 In contrast it has also been shown that the conversion of arginine to citrulline does not always lead to enhanced peptide MHC class II binding affinity. 15 We have also shown that HLA-DP4 transgenic mice make strong Th1 responses to human citrullinated vimentin and enolase peptides that do not cross react with wild type peptides. These responses also cross react with the homologous murine sequences suggesting a breaking of tolerance similar to the scenario that would be encountered in human patients.…”

“…To assess if responses to the citrullinated (cit) peptides could be restricted through HLA-DP4, the peptides were tested for binding to HLA-DP4. Binding was compared to a known HLA-DP4 restricted peptide from Hepatitis B surface antigen and two peptides from fibrinogen and collagen II that have been shown in the literature 15 not to bind to HLA-DP4. In our assay, the biotinylated Hepatitis B peptide showed good binding to HLA-DP4 whereas peptides from fibrinogen and collagen II demonstrated minimal binding over control (Figure 3(a)).…”

HLA associations in inflammatory arthritis: emerging mechanisms and clinical implications

“…Studies with other shared epitope-containing HLA-DR variants and more peptides have shown, however, that the amount of binding gained from P4-citrullination is variable, and in several cases no gain is apparent, suggesting that the magnitude of the effect is context-dependent 68,69 . Citrullination at other anchor or non-anchor residues usually has no effect on (or weakens) binding 68,70 . Importantly, to support a neoantigen hypothesis, it is not necessary to postulate that all P4-citrullination events result in improved binding to shared epitope-containing HLA-DR alleles -the loss of tolerance to a small number of neoantigens might be enough to explain an increased disease risk.…”

The interplay between citrullination and HLA-DRB1 polymorphism in shaping peptide binding hierarchies in rheumatoid arthritis

“…This is most likely attributable to polymorphic residues within the peptide-binding cleft of these HLA-SE allomorphs and is also reflected in the selection of naturally presented peptides by these different HLA-DR4 allomorphs. Moreover, the majority of these selfantigens were identified in studies utilizing either HLA-DRB1*04:01 RA patients PBMC or DR4/IE transgenic mice (Table 1), which may explain why certain citrullinated self-peptides bound better to HLA-DRB1*04:01 but not HLA-DRB1*04:04/*04:05 (35). Thus, there is a need for further epitope discovery in the context of these other HLA-DR4-SE allomorphs.…”