Proteomic analyses and identification of arginine methylated proteins differentially recognized by autosera from anti-Sm positive SLE patients

Background: antibodies against spliceosome sm proteins (anti-sm autoantibodies) are specific to the autoimmune disease systemic lupus erythematosus (sle). anti-sm autosera have been reported to specifically recognize sm d1 and d3 with symmetric di-methylarginines (sdma). we investigated if anti-sm sera from local sle patients can differentially recognize sm proteins or any other proteins due to their methylation states. results: we prepared hela cell proteins at normal or hypomethylation states (treated with an indirect methyltransferase inhibitor adenosine dialdehyde,adox). a few signals detected by the anti-sm positive sera from typical sle patients decreased consistently in the immunoblots of hypomethylated cell extracts. the differentially detected signals by one serum (sm1) were pinpointed by two-dimensional electrophoresis and identified by mass spectrometry. three identified proteins: splicing factor,proline- and glutamine-rich (sfpq),heterogeneous nuclear ribonucleoprotein d-like (hnrnp dl) and cellular nucleic acid binding protein (cnbp) are known to contain methylarginines in their glycine and arginine rich (gar) sequences. we showed that recombinant hnrnp dl and cnbp expressed in escherichia coli can be detected by all anti-sm positive sera we tested. as cnbp appeared to be differentially detected by the sle sera in the pilot study,differential recognition of arginine methylated cnbp protein by the anti-sm positive sera were further examined. hypomethylated flag-cnbp protein immunopurified from adox-treated hela cells was less recognized by sm1 compared to the cnbp protein expressed in untreated cells. two of 20 other anti-sm positive sera specifically differentiated the flag-cnbp protein expressed in hela cells due to the methylation. we also observed deferential recognition of methylated recombinant cnbp proteins expressed from e. coli by some of the autosera. conclusion: our study showed that hnrnp dl and cnbp are novel antigens for sle patients and the recognition of cnbp might be differentiated dependent on the level of arginine methylation. © 2013 chang et al.; licensee biomed central ltd.