Anti-U1C autoantibody interferes with the U1C-mediated splicing of both Ang1 and Tie2 genes in human pulmonary artery smooth muscle cells (159.10)

Abstract

Abstract Purpose: We previously showed that the frequency of the splicing variant of Angiopoietin-1 gene with nt805GGT insertion (Ang1/ins) was significantly increased in the Japanese patients with MCTD and those with PH (Shiozawa K et al., Arth. Rheum. 52: S283, ‘05). Because anti-U1RNP Ab is highly associated with the patients with MCTD, we investigated the correlation between anti-U1RNP Ab and the splicing of Ang1. Although we found that anti-U1C Ab specifically affects the splicing of Ang1 (Komai K et al., J. Immunol. 186: 44.3 (Abstr) '11), other genes also might be affected because U1RNP is the general splicing factor. We have now studied the contribution of anti-U1C Ab to the splicing of endothelial tyrosine kinase receptor Tie2 gene which codes the receptor of Ang1. Methods: Anti-U1C Abs were introduced into human pulmonary artery smooth muscle cells using PULS-in (Polyplus) with recombinant proteins of U1A, U1C or U1-70k respectively. After 24hrs, Tie2 mRNAs were obtained by RT-PCR. Results: Several Tie2 variants were induced with anti-U1C Ab only in the case with the introduction of U1C recombinant protein. Major products were 1100, 900, 600, 550, 450 and 400bp fragments. The 550 bp fragment was the Tie2-468 variant which was identified previously (Jin P et al., Arth. Res. & Ther. 10: R73, '08). Conclusions: Our results suggest that anti-U1C Ab interferes with the splicing of both Ang1 and Tie2 genes and induced variants may affect the pathogenesis of MCTD or those with PH.