Adaptor protein complex-1 (AP-1) and Golgi associated, γ-adaptin ear containing, Arf binding proteins (GGAs) are clathrin adaptors that regulate membrane trafficking between the trans-Golgi network (TGN) and endosomes. p56 is a clathrin adaptor accessory protein that may modulate the function of GGAs in mammalian cell lines. However, the precise relationship between p56 and the three GGAs (GGA1-3), as well as the physiological role of p56 in tissue cells, remain unknown. To this end, we generated an antibody against p56 and determined its cellular localization. In ARPE-19 cells and mouse embryonic fibroblasts, p56 was found to be localized as fine puncta in the TGN. Interestingly, the depletion of each clathrin adaptor by RNAi revealed that this localization was dependent on the expression of GGA1, but not that of GGA2, GGA3, or AP-1. Using immunohistofluorescence microscopy in the mouse central nervous system (CNS), p56 was clearly detected as scattered cytoplasmic puncta in spinal motor neurons, cerebellar Purkinje cells, and pyramidal neurons of the hippocampus and cerebral cortex. Moreover, double labeling with organelle markers revealed that the majority of these puncta were closely associated with the TGN; however, a small fraction was associated with endosomes or lysosomes in spinal motor neurons. Collectively, these results indicate a functional association of p56 with GGA1, suggesting an important role of p56 in larger CNS neurons.

中文翻译：

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GGA辅助蛋白p56在细胞系和中枢神经系统神经元中的胞内定位。

衔接蛋白复合物1（AP-1）和高尔基体相关，含γ-adaptin耳的Arf结合蛋白（GGA）是网格蛋白衔接体，调节跨高尔基体网络（TGN）和内体之间的膜运输。p56是一种网格蛋白衔接子辅助蛋白，可以调节哺乳动物细胞系中GGA的功能。然而，p56和三个GGA（GGA1-3）之间的确切关系以及p56在组织细胞中的生理作用仍然未知。为此，我们产生了针对p56的抗体并确定了其细胞定位。在ARPE-19细胞和小鼠胚胎成纤维细胞中，发现p56在TGN中定位为细小点。有趣的是，RNAi对每个网格蛋白衔接子的消耗表明该定位依赖于GGA1的表达，而不依赖于GGA2，GGA3或AP-1的表达。在小鼠中枢神经系统（CNS）中使用免疫组织荧光显微镜检查，p56被清楚地检测为脊髓运动神经元，小脑浦肯野细胞以及海马和大脑皮层锥体神经元中的散在的细胞质点。此外，用细胞器标记物进行的双重标记显示，大多数这些泪点都与TGN密切相关。然而，一小部分与脊髓运动神经元内体或溶酶体有关。总体而言，这些结果表明p56与GGA1的功能相关，表明p56在较大的CNS神经元中起重要作用。此外，用细胞器标记物进行的双重标记显示，大多数这些泪点都与TGN密切相关。然而，一小部分与脊髓运动神经元内体或溶酶体有关。总体而言，这些结果表明p56与GGA1的功能相关，表明p56在较大的CNS神经元中起重要作用。此外，用细胞器标记物进行的双重标记显示，大多数这些泪点都与TGN密切相关。然而，一小部分与脊髓运动神经元内体或溶酶体有关。总体而言，这些结果表明p56与GGA1的功能相关，表明p56在较大的CNS神经元中起重要作用。