Proinflammatory cytokines modulates expression of novel isoforms of PM/Scl-75 transcripts through alternative splicing: Pathogenic role in enhancing autoantigen immunogenicity in idiopathic inflammatory myositis

Abstract

Rationale In polymyositis-scleroderma (PM/Scl) overlap syndrome, autoantibodies to PM/Scl complex, a RNA exosome, have been frequently detected. Components of the RNA exosome include known autoantigens in patients with polymyositis, scleroderma or PM/Scl overlap syndrome, such as PM/Scl-75/Rrp45p and Rrp46p, etc. We showed that the upregulation of Rrp46p, a broadly immunogenic tumor antigen, is associated with its immunogenicity. We hypothesized that the proinflammatory cytokines interferon-γ (INF-γ), interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), whose expressions are often elevated in pathogenic muscle cells in patients with polymyositis, promote upregulation of autoantigens by post-transcriptional mechanisms in which a critical alternative splicing factor/splicing factor 2 (ASF/SF2) plays an important role. Methods To test this hypothesis, we stimulated differentiated mouse myotubes C2C12 with proinflammatory cytokines, and examined the expression of mRNA transcripts for autoantigens PM/Scl-75 by semi-quantitative PCR, and the expression of ASF/SF2 by Western blot. Results (1) Three alternatively spliced isoforms of PM/Scl-75 were identified through database mining. (2) Proinflammatory cytokines stimulate the expression of some alternatively-spliced isoforms of PM/Scl-75 while suppresssing others maximally at 6-24 hours. (3) Proinflammatory cytokines suppressed the expression of the ASF/SF2 maximally at 6 hours Conclusions We have demonstrated that proinflammatory cytokines can modulate expression of PM/Scl-75, by upregulation or downregulation of its isoforms through transcriptional and RNA splicing mechanisms that may be mediated through modulating ASF/SF2 expression. Alterations in the expression of different isoforms of autoantigens with various protein structures may result in generation of untolerized novel antigenic epitopes and changes in immunogenicity of autoantigens, which further leads to the loss of tolerance to these self-antigens.