Hata D, Kawakami T, Ishigami T, Kim K M, Heike T, Katamura K, Mayumi M, Mikawa H
Department of Pediatrics, Faculty of Medicine, Kyoto University, Japan.
Int Immunol. 1992 Jul;4(7):797-804. doi: 10.1093/intimm/4.7.797.
We investigated tyrosine phosphorylation and structural properties of the IgM-associated molecules in comparison with IgD-associated molecules in a recently established human surface IgM+, IgD+ B lymphoma cell line, B104, the growth of which was irreversibly inhibited by anti-IgM mAbs but not by anti-IgD mAbs. Tyrosine kinase activity and tyrosine phosphorylated proteins were detected in anti-IgM and anti-IgD immunoprecipitates from digitonin lysates of B104 cells with the use of an in vitro kinase assay followed by a re-immunoprecipitation experiment with anti-phosphotyrosine mAbs. Tyrosine phosphorylated proteins of 74, 58-44, 41, and 39 kDa were detected in anti-IgM immunoprecipitates, whereas tyrosine phosphorylated proteins of 74, 58-44, and 39 kDa, but not 41 kDa, were detected in anti-IgD immunoprecipitates. Crosslinking of surface IgM and surface IgD stimulated rapid tyrosine phosphorylation of different sets of proteins which included tyrosine-phosphorylated proteins of the same or similar molecular weights as those detected in the anti-IgM and anti-IgD immunoprecipitates respectively. After deglycosylation by N-glycosidase, both the IgM- and IgD-associated phosphoproteins (pp58-pp39) gave rise to the same three bands of 29, 27, and 26 kDa. Proteolytic peptide mapping of these three deglycosylated proteins showed that the primary structures of the IgM- and IgD-associated molecules are identical, suggesting that the IgM- and IgD-associated phosphoproteins (pp58-pp39) are the products of the same or closely related genes. One of the products, pp41, may be associated with IgM, but not with IgD, although the same gene product may be associated with IgD in a different glycosylation pattern.
我们在最近建立的人表面IgM⁺、IgD⁺ B淋巴瘤细胞系B104中，研究了与IgM相关分子的酪氨酸磷酸化和结构特性，并与IgD相关分子进行比较。该细胞系的生长被抗IgM单克隆抗体不可逆地抑制，而不被抗IgD单克隆抗体抑制。利用体外激酶测定法，随后用抗磷酸酪氨酸单克隆抗体进行再免疫沉淀实验，在B104细胞的洋地黄皂苷裂解物的抗IgM和抗IgD免疫沉淀物中检测到酪氨酸激酶活性和酪氨酸磷酸化蛋白。在抗IgM免疫沉淀物中检测到74、58 - 44、41和39 kDa的酪氨酸磷酸化蛋白，而在抗IgD免疫沉淀物中检测到74、58 - 44和39 kDa的酪氨酸磷酸化蛋白，但未检测到41 kDa的。表面IgM和表面IgD的交联刺激了不同组蛋白的快速酪氨酸磷酸化，这些组蛋白分别包括与在抗IgM和抗IgD免疫沉淀物中检测到的分子量相同或相似的酪氨酸磷酸化蛋白。用N -糖苷酶去糖基化后，与IgM和IgD相关的磷蛋白（pp58 - pp39）都产生了29、27和26 kDa的相同三条带。这三种去糖基化蛋白的蛋白水解肽图谱显示IgM和IgD相关分子的一级结构相同，表明与IgM和IgD相关的磷蛋白（pp58 - pp39）是相同或密切相关基因的产物。其中一种产物pp41可能与IgM相关，但与IgD无关，尽管相同的基因产物可能以不同的糖基化模式与IgD相关。
