Receptor-Mediated Endocytosis of an Extracellular Steroid-Binding Protein (TeBG) in MCF-7 Human Breast Cancer Cells*

作者：Catarina Segreti Porto, Glen L. Gunsalus, C. Wayne Bardin, David M. Phillips, Neal A. Musto · 发表于：Endocrinology · 年份：1991 · DOI：10.1210/endo-129-1-436 · 被引用次数：46 · 研究领域：Lipid Membrane Structure and Behavior、Receptor Mechanisms and Signaling、Hypothalamic control of reproductive hormones

Previous studies suggested that an extracellular steroid-binding protein, testosterone-estradiol-binding globulin (TeBG), can enter a variety of cells. Experiments were conducted to determine whether uptake of TeBG occurs by nonspecific fluid phase endocytosis or via a specific receptor-mediated process. In human breast carcinoma cells (MCF-7) maintained on serum-free medium, exposure to radiolabeled TeBG resulted in cellular uptake, which reached a plateau by 6 h and could be inhibited 80% by competition with unlabeled TeBG. Uptake was temperature dependent with cell-associated radioactivity at 37 C being 1.6-fold higher than at 4 C. Subsequent exposure of cells to pronase resulted in release of the cell-associated TeBG by 88% and 44% at 4 C and 37 C, respectively. After transfer to media devoid of TeBG, approximately 35% of cell-associated radioactivity was release into the medium at 37 C; it was not possible to distinguish whether this was released from the cell surface or from inside the cell. Investigation of the localization of TeBG-gold complexes by electron microscopy revealed that TeBG first binds to the plasmalemma. Within 15 min label appears in receptosomes, which fuse to form multivesicular endosomes. By 1 h all label is observed in multivesicular endosomes and lysosomes, most of which are in the Golgi zone. Localization of the internalized radioactivity using classical cell fractionation techniques showed it appears in a symmetrical band exhibiting the same buoy...