Wada N, Kimura I, Tanaka H, Ide H, Nohno T
Department of Molecular Biology, Kawasaki Medical School, 577 Matsushima, Kurashiki 7, 01-0192, Japan.
Dev Biol. 1998 Oct 15;202(2):244-52. doi: 10.1006/dbio.1998.9013.
Although regional differences in mesenchymal cell affinity in the limb bud represent positional identity, the molecular basis for cell affinity is poorly understood. We found that treatment of the cell surface with bacterial phosphatidylinositol-specific phospholipase C (PI-PLC) could change cell affinity in culture. When PI-PLC was added to the culture medium, segregation of the progress zone (PZ) cells from different stage limb buds was inhibited. Similarly, sorting out of the cells from different positions along the proximodistal (PD) axis of the same stage limb buds was disturbed. Since PI-PLC can remove glycosylphosphatidylinositol (GPI)-anchored membrane bound proteins from the cell surface, the GPI-anchored cell surface proteins may be involved in sorting out. To define the GPI-anchored molecules that determine the segregation of limb mesenchymal cells, we examined the effect of neutralizing antibody on the EphA4 receptor that binds to GPI-anchored cell surface ligands, called ephrin-A. Sorting out of the PZ cells at different stages could be inhibited by the neutralizing antibody to EphA4. These results suggest that EphA4 and its GPI-anchored ligands are, at least in part, involved in sorting out of limb mesenchymal cells with different proximal-distal positional values, and that GPI-anchored cell surface proteins play important roles in determining cell affinity in the limb bud.
尽管肢芽中间充质细胞亲和力的区域差异代表了位置身份，但细胞亲和力的分子基础仍知之甚少。我们发现，用细菌磷脂酰肌醇特异性磷脂酶C（PI-PLC）处理细胞表面可改变培养中的细胞亲和力。当将PI-PLC添加到培养基中时，不同阶段肢芽的进展区（PZ）细胞的分离受到抑制。同样，沿同一阶段肢芽近端-远端（PD）轴不同位置的细胞分选也受到干扰。由于PI-PLC可以从细胞表面去除糖基磷脂酰肌醇（GPI）锚定的膜结合蛋白，因此GPI锚定的细胞表面蛋白可能参与分选。为了确定决定肢体间充质细胞分离的GPI锚定分子，我们研究了中和抗体对与GPI锚定的细胞表面配体（称为ephrin-A）结合的EphA4受体的作用。针对EphA4的中和抗体可以抑制不同阶段PZ细胞的分选。这些结果表明，EphA4及其GPI锚定配体至少部分参与了具有不同近端-远端位置值的肢体间充质细胞的分选，并且GPI锚定的细胞表面蛋白在决定肢芽中的细胞亲和力方面发挥着重要作用。
