Miñana R, Duran J M, Tomas M, Renau-Piqueras J, Guerri C
Instituto de Investigaciones Citológicas (FVIB), Amadeo de Saboya 4, 46010-Valencia, Spain.
Eur J Neurosci. 2001 Feb;13(4):749-56. doi: 10.1046/j.0953-816x.2000.01439.x.
Neural cell adhesion molecule (NCAM) constitutes a group of cell surface glycoproteins that regulate cell-cell interactions in the developing and adult brain. Endocytosis is a mechanism which dynamically controls the amount of cell surface NCAM expression and may involve the rapid changes occurring in NCAM expression under certain physiological or pathological conditions. However, the endocytic pathway of NCAM is presently unknown. Using astrocytes in culture and immunofluorescence we show that NCAM is internalized and that the immunolabelling presents a high degree of colocalization with clathrin, alpha-adaptin and transferrin, suggesting that NCAM is endocytosed by a clathrin-dependent pathway. Potassium depletion which disrupts clathrin-mediated endocytosis, inhibited internalization of NCAM. Electron microscopy and immunogold studies also demonstrate that the surface of clathrin-coated vesicles are also immunolabelled for both alpha-adaptin and PSA-NCAM, the highly sialylated isoform of NCAM. Furthermore, immunoprecipation studies demonstrate that NCAM is associated with both clathrin and alpha-adaptin, a component of adaptor complex AP-2, in brain, neurons and astrocytes. These findings indicate that NCAM is mainly endocytosed via clathrin-coated vesicles, suggesting a possible mechanism that may contribute to the rapid changes in NCAM expression at the cell surface.
神经细胞黏附分子（NCAM）构成了一组细胞表面糖蛋白，可调节发育中和成体大脑中的细胞间相互作用。内吞作用是一种动态控制细胞表面NCAM表达量的机制，可能涉及在某些生理或病理条件下NCAM表达的快速变化。然而，目前尚不清楚NCAM的内吞途径。利用培养的星形胶质细胞和免疫荧光技术，我们发现NCAM被内化，并且免疫标记与网格蛋白、α-衔接蛋白和转铁蛋白呈现高度共定位，这表明NCAM通过网格蛋白依赖途径被内吞。破坏网格蛋白介导的内吞作用的钾离子耗竭抑制了NCAM的内化。电子显微镜和免疫金研究还表明，网格蛋白包被小泡的表面也被α-衔接蛋白和PSA-NCAM（NCAM的高度唾液酸化异构体）免疫标记。此外，免疫沉淀研究表明，在脑、神经元和星形胶质细胞中，NCAM与网格蛋白和衔接蛋白复合体AP-2的一个组分α-衔接蛋白相关联。这些发现表明，NCAM主要通过网格蛋白包被小泡被内吞，提示了一种可能有助于细胞表面NCAM表达快速变化的机制。
