Zhao J, Zhou Q, Wiedmer T, Sims P J
Blood Center of Southeastern Wisconsin, Milwaukee, Wisconsin 53201, USA.
J Biol Chem. 1998 Mar 20;273(12):6603-6. doi: 10.1074/jbc.273.12.6603.
We recently identified a 35-kDa erythrocyte membrane protein, phospholipid scramblase, that promotes Ca2+-dependent transbilayer movement of phosphatidylserine (PS) and other phospholipids (PL) in reconstituted proteoliposomes (Zhou, Q., Zhao, J., Stout, J. G., Luhm, R. A., Wiedmer, T., and Sims, P. J. (1997) J. Biol. Chem. 272, 18240-18244). To determine whether this same protein is responsible for the rapid movement of PS from inner-to-outer plasma membrane leaflets in other cells exposed to elevated cytosolic calcium concentration ([Ca2+]c), we analyzed how induced movement of PS to the cell surface related to expression of PL scramblase. Exposure to Ca2+ ionophore A23187 resulted in rapid PS exposure in those cell lines constitutively high in PL scramblase (HEL, Epstein-Barr virus-transformed B-lymphocytes, and Jurkat), whereas this response was markedly attenuated in cells expressing low amounts of this protein (Raji, HL60, and Dami). To confirm this apparent correlation between PL scramblase expression and PS egress at elevated [Ca2+]c, Raji cells were transfected with PL scramblase cDNA in pEGFP-C2, and stable transformants expressing various amounts of GFP-PL scramblase fusion protein were obtained. Clones expressing GFP-PL scramblase showed distinctly plasma membrane-localized fluorescence. When compared either with untransfected Raji cells or with transformants expressing GFP alone, clones expressing GFP-PL scramblase fusion protein showed increased exposure of PS at the cell surface in response to elevated [Ca2+]c, accompanied by increased expression of membrane catalytic function for the prothrombinase enzyme complex. These data indicate that transfection with PL scramblase cDNA promotes movement of PS to cell surfaces and suggest that this protein normally mediates redistribution of plasma membrane phospholipids in activated, injured, or apoptotic cells.
我们最近鉴定出一种35 kDa的红细胞膜蛋白——磷脂翻转酶，它能促进磷脂酰丝氨酸（PS）和其他磷脂（PL）在重组蛋白脂质体中依赖Ca2+的跨膜运动（周，Q.，赵，J.，斯托特，J.G.，卢姆，R.A.，维德默，T.，以及西姆斯，P.J.（1997年）《生物化学杂志》272卷，18240 - 18244页）。为了确定在其他细胞中，当胞质钙浓度（[Ca2+]c）升高时，是否是同一种蛋白负责PS从质膜内小叶快速移动到外小叶，我们分析了PS向细胞表面的诱导移动与PL翻转酶表达之间的关系。暴露于Ca2+离子载体A23187会导致那些PL翻转酶组成性高表达的细胞系（HEL、爱泼斯坦 - 巴尔病毒转化的B淋巴细胞和Jurkat细胞）中PS快速暴露于细胞表面，而在表达该蛋白量低的细胞（Raji、HL60和Dami细胞）中，这种反应明显减弱。为了证实PL翻转酶表达与[Ca2+]c升高时PS外流之间的这种明显相关性，用pEGFP - C2中的PL翻转酶cDNA转染Raji细胞，并获得了表达不同量GFP - PL翻转酶融合蛋白的稳定转化体。表达GFP - PL翻转酶的克隆显示出明显的质膜定位荧光。与未转染的Raji细胞或单独表达GFP的转化体相比，表达GFP - PL翻转酶融合蛋白的克隆在[Ca2+]c升高时，细胞表面PS暴露增加，同时凝血酶原酶复合物的膜催化功能表达增加。这些数据表明，用PL翻转酶cDNA转染可促进PS向细胞表面移动，并提示该蛋白通常介导活化、损伤或凋亡细胞中质膜磷脂的重新分布。
