Smythe E, Smith P D, Jacob S M, Theobald J, Moss S E
Department of Physiology, University College London, United Kingdom.
J Cell Biol. 1994 Feb;124(3):301-6. doi: 10.1083/jcb.124.3.301.
Annexin VI is one of a family of calcium-dependent phospholipid-binding proteins. Although the function of this protein is not known, various physiological roles have been proposed, including a role in the budding of clathrin-coated pits (Lin et al., 1992. Cell. 70:283-291.). In this study we have investigated a possible endocytotic role for annexin VI in intact cells, using the human squamous carcinoma cell line A431, and report that these cells do not express endogenous annexin VI, as judged by Western and Northern blotting and PCR/Southern blotting. To examine whether endocytosis might in some way be either facilitated or inhibited by the presence of annexin VI, a series of A431 clones were isolated in which annexin VI expression was achieved by stable transfection. These cells expressed annexin VI at similar levels to other human cell types. Using assays for endocytosis and recycling of the transferrin receptor, we report that each of these cellular processes occurs with identical kinetics in both transfected and wild-type A431 cells. In addition, purified annexin VI failed to support the scission of coated pits in permeabilized A431 cells. We conclude that annexin VI is not an essential component of the endocytic pathway, and that in A431 cells, annexin VI fails to exert any influence on internalization and recycling of the transferrin receptor.
膜联蛋白VI是一类钙依赖性磷脂结合蛋白家族的成员之一。尽管该蛋白的功能尚不清楚，但已提出了多种生理作用，包括在网格蛋白包被小窝出芽过程中发挥作用（Lin等人，1992年。《细胞》。70:283 - 291）。在本研究中，我们使用人鳞状癌细胞系A431，研究了膜联蛋白VI在完整细胞中可能的内吞作用，并报告这些细胞不表达内源性膜联蛋白VI，这通过蛋白质免疫印迹、Northern印迹以及PCR/ Southern印迹分析得以判断。为了研究膜联蛋白VI的存在是否可能以某种方式促进或抑制内吞作用，我们分离了一系列A431克隆，通过稳定转染使它们表达膜联蛋白VI。这些细胞表达的膜联蛋白VI水平与其他人类细胞类型相似。通过对转铁蛋白受体的内吞和循环分析，我们报告在转染的和野生型A431细胞中，这些细胞过程中的每一个都以相同的动力学发生。此外，纯化的膜联蛋白VI未能支持通透化的A431细胞中包被小窝的切割。我们得出结论，膜联蛋白VI不是内吞途径的必需成分，并且在A431细胞中，膜联蛋白VI对转铁蛋白受体的内化和循环没有任何影响。
