Nishida Tomoki, Arii Tatsuo, Takaoka Akio, Yoshimura Ryoichi, Endo Yasuhisa
Division of Applied Biology, Kyoto Institute of Technology, Sakyoku, Kyoto 606-8585, Japan.
Acta Histochem Cytochem. 2007 Jul 3;40(3):93-9. doi: 10.1267/ahc.07009.
Signaling of nerve growth factor (NGF) and its receptor (TrkA) promotes neuronal differentiation, synapse formation and survival. It has been known that the complex of NGF and TrkA is internalized into the cytoplasm and transported for further signal transduction, but the ultrastructural information of this process is virtually unknown. In order to clarify the relationship between the internalization of TrkA and the membrane-associated proteins (caveolin and clathrin), the localization and three-dimensional structures of those proteins were examined with computer tomography of high voltage electron microscopy in PC12 cells. TrkA immunoreactivity was found only at definite areas in the plasma membrane, as ring and cluster structures. Its 3D image indicated that those cluster structures contained small pits, which did not appear to be typical caveolae in size and shape. 3D images of clathrin and caveolin-1 immunoreactivities indicated that the formation of those small pits was associated with clathrin, but not with caveolin-1. Caveolin-1 immunoreactivity was found as a mesh-like structure just beneath the plasma membrane. These results suggest that clathrin rather than caveolin is mainly involved in the process of TrkA internalization, at least in differentiated PC12 cells.
神经生长因子（NGF）及其受体（TrkA）的信号传导促进神经元分化、突触形成和存活。已知NGF与TrkA的复合物被内化到细胞质中并运输以进行进一步的信号转导，但这一过程的超微结构信息实际上并不清楚。为了阐明TrkA内化与膜相关蛋白（小窝蛋白和网格蛋白）之间的关系，利用高压电子显微镜计算机断层扫描技术检测了PC12细胞中这些蛋白的定位和三维结构。仅在质膜的特定区域发现TrkA免疫反应性，呈环状和簇状结构。其三维图像表明，这些簇状结构包含小凹，其大小和形状似乎不是典型的小窝。网格蛋白和小窝蛋白-1免疫反应性的三维图像表明，这些小凹的形成与网格蛋白有关，而与小窝蛋白-1无关。小窝蛋白-1免疫反应性表现为质膜下方的网状结构。这些结果表明，至少在分化的PC12细胞中，主要参与TrkA内化过程的是网格蛋白而非小窝蛋白。
