Molecular cloning of a cDNA clone encoding a phosphoprotein component related to the Ig receptor-mediated signal transduction

S. Inui, K. Kuwahara, J. Mizutani, K. Maeda, T. Kawai, H. Nakayasu, N. Sakaguchi

研究成果: Article査読

65 被引用数 (Scopus)

抄録

We have shown previously that a 52-kDa phosphoprotein (p52) co- precipitated with Ig receptor (IgR)-associated MB-1 protein was inducibly phosphorylated by the stimulation with 12-O-tetradecanoyl-phorbol-13-acetate. By immunizing the 52-kDa protein co-precipitated with MB-1, we prepared a mAb (19-14), which can immunoprecipitate the p52 and the associated kinase molecule. Immune complex kinase assay with the 19-14 mAb showed that the p52 is associated with a novel kinase molecule and is involved in IgR-mediated signal transduction. Here we isolated a cDNA clone (α4) from a murine bone marrow cDNA library in λgt-11 vector by the 19-14 mAb. The α4 cDNA encodes a novel protein of 340 amino acids with multiple potential phosphorylation sites. The 1.4-kb α4 mRNA is expressed in various cells including cell lines of B lineage, T lineage, monocytic, fibroblast, and normal organs such as liver, spleen, thymus, and brain. To study the molecule encoded by the α4 cDNA, we produced a chimeric protein of the glutathione S transferase (GST)- α4 in pGEX-3X expression vector. The 19-14 mAb binds to the GST-α4 fusion protein. Rabbit anti-α4 Ab prepared by immunizing the GST-α4 fusion protein immunoprecipitated a 52-kDa phosphoprotein from WEHI-231 cells. The 52-kDa phosphoprotein immunoprecipitated by the anti-α4 Ab is tightly associated with kinase activity that is similarly observed with the p52 protein reported previously. Moreover, the 52-kDa phosphoprotein immunoprecipitated with the anti-α4 Ab is identical to the p52 by the protein comparison on non- equilibrium pH gradient gel electrophoresis/SDS-PAGE, isoelectric focusing/SDS-PAGE, and V8 protease mapping. The 52-kDa phosphoprotein is associated with functional components that are inducibly phosphorylated by anti-IgM stimulation. These results suggested that the α4 gene-encoded molecule is functionally involved in the IgR-mediated signal transduction in B cells.

本文言語English
ページ(範囲)2714-2723
ページ数10
ジャーナルJournal of Immunology
154
6
出版ステータスPublished - 01-01-1995
外部発表はい

All Science Journal Classification (ASJC) codes

  • 免疫アレルギー学
  • 免疫学

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