Role of cell adhesion molecules during net formation in Hydorodictyon reticulatum

一种单克隆抗体，提出了对游动孢子的匀浆识别细胞粘附分子定位到游动孢子之间的接触部位。该分子在游动孢子形成网时持续表达，游动孢子间粘附完成后表达量下降。通过间接免疫荧光显微镜分析了该分子在网形成过程中的亚细胞定位。球形游动孢子的周边区域和多角形游动孢子的接触区域被明亮地染色。多面体游动孢子运动停止后，该分子立即位于两个细胞的紧密贴壁表面上。为了检查分子的精确空间分布，用该单克隆抗体和抗鸡β-微管蛋白的单克隆抗体对游动孢子和网细胞进行双重染色。该分子在多面体游动孢子和网细胞的接触区与微管束紧密共定位。秋水仙素处理的游动孢子不能形成六边形的网状结构，游动孢子周围区域染色均匀。免疫电子显微镜显示存在的金颗粒上的高尔基堆和质膜的多面体游动孢子和接触区的网细胞。我用这种单克隆抗体处理了分离的游动孢子。用抗体处理抑制六边形网的形成。这些结果表明，本地化的分子参与游动孢子的接触位点的确定和微管束参与本地化的分子在六边形的网络形成。

A monoclonal antibody that was raised against a homogenate of zoospores recognized cell adhesion molecules localized to the contact sites between zoospores. The molecule was constantly expressed in zoospores during net formation and decreased after the completion of the adhesion between zoospores. The subcellular localization of the molecule during net formation was analyzed by indirect immunofluorescence microscopy. The peripheral regions of the spherical zoospores and the contact regions of the polyhedral zoospores were stained brightly. Immediately after cessation of the movement of polyhedral zoospores, the molecule was located on the closely apposed surfaces of two cells. In order to examine the precise spatial distribution of the molecule, zoospores and net cells were double-stained with this monoclonal antibody and a monoclonal antibody against chicken β-tubulin. The molecule closely colocalized with the microtubule bundles at the contact regions of the polyhedral zoospores and the net cells. Colchicine-treated zoospores could not form hexagonal nets, then the peripheral regions of zoospores stained uniformly. Immunoelectron microscopy revealed the presence of gold particles on a Golgi stack and the plasma membrane in polyhedral zoospores and the contact regions of net cells. I treated isolated zoospores with this monoclonal antibody. Treatment with the antibody inhibited formation of hexagonal nets. These results suggest that localization of the molecule participate in the determination of the contact sites of zoospores and that the microtubule bundles are involved in the localization of the molecule during hexagonal net formation.