Stephan V, Guo N, Ginsburg V, Siraganian R P
Clinical Immunology Section, National Institute of Dental Research, National Institutes of Health, Bethesda, MD 20892.
J Immunol. 1991 Jun 15;146(12):4271-7.
In previous studies, mAb AA4 inhibited IgE binding, induced rapid morphologic changes, and blocked histamine release in rat basophilic leukemia (RBL-2H3) cells. It bound to two novel derivatives of ganglioside GD1b (Ag I and Ag II) that appear to be present only in rat mast cells. The present study demonstrates the importance of gangliosides Ag I and Ag II for binding of mAb AA4 to intact cells. We also investigated the presence of gangliosides Ag I and Ag II and proteins immunoprecipitated with mAb AA4 in the parental and four variant cell lines. In comparison with the parental RBL-2H3, two variant cell lines had very low (0.5% and 2.0%) and two others had intermediate levels (9% and 18%) of 125I-AA4 binding. mAb AA4 inhibited 125I-IgE binding to the parental RBL-2H3 cells and to only one variant with intermediate amounts of gangliosides Ag I and Ag II. Therefore, there are variations in the proximity of these gangliosides to the high affinity IgE receptor (Fc epsilon RI) among different cell lines. mAb AA4 immunoprecipitated proteins of 50 to 60, 120, and 135 kDa from 125I-surface labeled cells. These were different from the subunits of Fc epsilon RI. The amount of gangliosides Ag I and Ag II in cell extracts correlated with the number of mAb AA4 binding sites on the cell surface and with the quantity of proteins precipitated from the different cell lines. Thus, these membrane proteins appear to be associated with gangliosides Ag I and Ag II. The binding of mAb AA4 to the surface gangliosides could induce intracellular changes through transmembrane signaling by these proteins.
在先前的研究中，单克隆抗体AA4可抑制IgE结合，诱导快速的形态学变化，并阻断大鼠嗜碱性白血病（RBL - 2H3）细胞中的组胺释放。它与神经节苷脂GD1b的两种新衍生物（抗原I和抗原II）结合，这两种衍生物似乎仅存在于大鼠肥大细胞中。本研究证明了神经节苷脂抗原I和抗原II对于单克隆抗体AA4与完整细胞结合的重要性。我们还研究了亲代细胞系和四个变异细胞系中神经节苷脂抗原I和抗原II以及用单克隆抗体AA4免疫沉淀的蛋白质的存在情况。与亲代RBL - 2H3相比，两个变异细胞系的125I - AA4结合水平非常低（分别为0.5%和2.0%），另外两个变异细胞系的结合水平处于中等（分别为9%和18%）。单克隆抗体AA4可抑制125I - IgE与亲代RBL - 2H3细胞以及仅一种含有中等量神经节苷脂抗原I和抗原II的变异细胞系的结合。因此，不同细胞系中这些神经节苷脂与高亲和力IgE受体（FcεRI）的接近程度存在差异。单克隆抗体AA4从125I表面标记的细胞中免疫沉淀出50至60 kDa、120 kDa和135 kDa的蛋白质。这些蛋白质与FcεRI的亚基不同。细胞提取物中神经节苷脂抗原I和抗原II的量与细胞表面单克隆抗体AA4结合位点的数量以及从不同细胞系中沉淀出的蛋白质数量相关。因此，这些膜蛋白似乎与神经节苷脂抗原I和抗原II相关。单克隆抗体AA4与表面神经节苷脂的结合可通过这些蛋白质的跨膜信号传导诱导细胞内变化。
