Muller L, Picart R, Barret A, Bockaert J, Homburger V, Tougard C
Groupe de Biologie de la Cellule Neuroendocrine, Collège de France, CNRS URA 1115.
Mol Cell Neurosci. 1994 Dec;5(6):556-66. doi: 10.1006/mcne.1994.1068.
The subcellular distribution of multiple subunits of heterotrimeric GTP-binding proteins has been investigated in rat anterior pituitary cells in primary culture, and more precisely in prolactin cells, by immunocytochemistry and subcellular fractionation followed by immunoblotting or ADP ribosylation, using polyclonal affinity-purified antibodies directed against Gi3 alpha, Gs alpha, Go1 alpha, Go2 alpha, and G beta. As expected, all these subunits were detected on the plasma membrane. They were, however, also detected on the membrane of several intracellular compartments involved in the secretory pathway, particularly on the secretory granule membrane. Differences appeared between the precise subcellular distribution and the local concentration of each subunit. The main subunits present on the secretory granule membrane were Gi3 alpha and Gs alpha. Go1 alpha, Go2 alpha, and G beta were detected, to a lesser extent, on parts of the membrane of a few secretory granules located near the plasma membrane. Domains of the rough endoplasmic reticulum cisternae were immunolabeled with anti-Gs alpha and anti-Go1 alpha. In the Golgi zone, the membrane of some vesicles was stained only with anti-Gs alpha and anti-Go2 alpha. The association of this set of heterotrimeric G protein subunits on the membrane of the secretory granules suggests that these subunits could be involved in the regulation of formation, storage, targeting, and/or exocytosis of these organelles.
通过免疫细胞化学和亚细胞分级分离，随后进行免疫印迹或 ADP 核糖基化，使用针对 Gi3α、Gsα、Go1α、Go2α 和 Gβ 的多克隆亲和纯化抗体，研究了原代培养的大鼠垂体前叶细胞，更确切地说是催乳素细胞中异三聚体 GTP 结合蛋白多个亚基的亚细胞分布。正如预期的那样，所有这些亚基都在质膜上被检测到。然而，它们也在分泌途径中涉及的几个细胞内区室的膜上被检测到，特别是在分泌颗粒膜上。每个亚基的精确亚细胞分布和局部浓度之间出现了差异。分泌颗粒膜上存在的主要亚基是 Gi3α 和 Gsα。在靠近质膜的少数分泌颗粒膜的部分上，较少程度地检测到 Go1α、Go2α 和 Gβ。粗面内质网池的区域用抗 Gsα 和抗 Go1α 进行了免疫标记。在高尔基体区域，一些小泡的膜仅用抗 Gsα 和抗 Go2α 染色。这组异三聚体 G 蛋白亚基在分泌颗粒膜上的结合表明，这些亚基可能参与这些细胞器的形成、储存、靶向和/或胞吐作用的调节。
