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Altered posttranslational modification on U 1 small nuclear ribonucleoprotein 68k in systemic autoimmune diseases detected by 2D Western blot
Author(s) -
Nagai Kouhei,
Arito Mitsumi,
Takakuwa Yukiko,
Ooka Seido,
Sato Toshiyuki,
Kurokawa Manae S.,
Okamoto Kazuki,
Uchida Teisuke,
Suematsu Naoya,
Kato Tomohiro
Publication year - 2012
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201200058
Subject(s) - autoantibody , extractable nuclear antigens , western blot , mixed connective tissue disease , peripheral blood mononuclear cell , gene isoform , antibody , ribonucleoprotein , microbiology and biotechnology , heterogeneous nuclear ribonucleoprotein , chemistry , antigen , immunology , anti nuclear antibody , medicine , biology , gene , biochemistry , rna , in vitro
Anti‐ribonucleoprotein (anti‐ RNP ) antibodies are one of the representative autoantibodies detectable in patients with systemic lupus erythematosus ( SLE ) and mixed connective tissue disease ( MCTD ). Generally, posttranslational modifications ( PTM s) on autoantigens are proposed to be involved in the production of autoantibodies. In this study, we tried to detect the alteration in PTM s on a U 1 small nuclear RNP 68k subunit ( U 1‐68k), a major antigen of anti‐ RNP antibodies. Peripheral blood mononuclear cells ( PBMC s) were obtained from patients with MCTD , SLE , and rheumatoid arthritis ( RA ), and from healthy donors. U 1‐68ks in the PBMC s were detected by 2D Western blot (WB), where extracted nuclear proteins were separated by 2 DE , followed by the detection of U 1‐68k using WB. More than 20 PTM isoforms were detected with different molecular weights of 65.0 , 66.5, and 68.0kDa, and different p I s between 6.0 and 8.5. Importantly, the relative intensity of the spot with 66.5 kDa and p I 7.5 was significantly increased in the MCTD and SLE groups compared to the RA and healthy groups. Further, this U 1‐68k isoform, in particular, in its RS domain, was found to have significantly decreased phosphorylation compared to the other isoforms. The PTM alternation may be one of the steps to generate the anti‐ RNP antibodies.