Systemic Lupus Erythematosus Patients Contain Significantly Less IgM against Mono-Methylated Lysine than Healthy Subjects

Abstract

Post-translational modifications on proteins are important in biological processes but may create neo-epitopes that induce autoimmune responses. In this study, we measured the serum IgG and IgM response to a set of non-modified or acetyl- and methyl-modified peptides corresponding to residues 1–19 of the histone 3 N-terminal tail in systemic lupus erythematosus (SLE) patients and healthy subjects. Our results indicated that the SLE patients and healthy subjects produced antibodies (Abs) to the peptides, but the two groups had different Ab isotype and epitope preferences. Abs to the non-modified form, H31–19, were of the IgG isotype and produced by SLE patients. They could not recognize the scrambled H31–19, which contained the same amino acid composition but a different sequence as H31–19. In comparison, healthy subjects in general did not produce IgG against H31–19. However, about 70% of the healthy subjects produced IgM Abs against mono-methylated K9 of H31–19 (H31–19K9me). Our further studies revealed that ε-amine mono-methylated lysine could completely inhibit the IgM binding to H31–19K9me, but lysine had no inhibitory effect. In addition, the IgM Abs could bind peptides containing a mono-methylated lysine residue but with totally different sequences. Thus, mono-methylated lysine was the sole epitope for the IgM. Interestingly, SLE patients had much lower levels of this type of IgM. There was no obvious correlation between the IgM levels and disease activity and the decreased IgM was unlikely caused by medical treatments.We also found that the IgM Abs were not polyreactive to dsDNA, ssDNA, lipopolysaccharide (LPS) or insulin and they did not exist in umbilical cord serum, implying that they were not natural Abs. The IgM Abs against mono-methylated lysine are present in healthy subjects but are significantly lower in SLE patients, suggesting a distinct origin of production and special physiological functions.

Autoantibodies against mono- and tri-methylated lysine display similar but also distinctive characteristics

“…The levels of total IgM in SLE patients were also lower than those in healthy subjects ( Fig 6C ), whereas the levels of total IgG in SLE and other rheumatic patients were higher than those in healthy subjects ( Fig 6D ). As previously reported [ 12 ], there was no correlation between the levels of anti-Kme IgM and disease activity of SLE ( P = 0.5774, Fig 6E ). There was also no correlation between the levels of anti-Kme3 IgM and disease activity of SLE ( P = 0.3276, Fig 6F ).…”

“…The purified antibodies were highly specific. We previously showed that IgM anti-Kme reacted to peptides that contained mono-methylated lysine and did not react to peptides that had no methylation or had di- or tri-methylated lysine [ 12 ]. To determine whether the IgM reacted with methyl groups on other residues, we tested its reactivity to methylated DNA and mono-methylated arginine that was on a peptide corresponding to the 1–19 N-terminal residues of histone H3 (H3 1-19 R2me, Table 1 ).…”

“…We previously reported that serum levels of IgM anti-Kme were significantly lower in SLE patients than healthy subjects [ 12 ]. To determine whether IgM anti-Kme levels were also low in other rheumatic diseases, we tested sera from patients with RA, SS and AS.…”

“…In previous work, we identified IgM autoantibodies that reacted to a very small epitope with only one amino acid, i.e., mono-methylated lysine (Kme) [ 12 ]. The antibodies were not polyreactive.…”

Autoantibodies against Modified Histone Peptides in SLE Patients Are Associated with Disease Activity and Lupus Nephritis

“…The unexpected lack of specificity for anti-H3p/H3p me reactivity in our assay might be due to a decreased accessibility within the circulating nucleosome/apoptotic microparticle, for which we found evidence in a previous study [6]. Another explanation might be the reactivity against methylated residues in plasmas from healthy individuals, which has been described before for IgM [32]. Importantly, we demonstrated that the modified peptides H4p ac and H2Bp ac show an increased specificity and sensitivity compared to their unmodified counterpart.…”