Lublin D M, Coyne K E
Department of Pathology, Washington University School of Medicine, St. Louis, Missouri 63110.
J Exp Med. 1991 Jul 1;174(1):35-44. doi: 10.1084/jem.174.1.35.
Decay-accelerating factor (DAF) is a glycosyl-phosphatidylinositol (GPI)-anchored membrane protein that protects cells from complement-mediated damage by regulation of the C3 convertase. To investigate the role of the GPI anchor in the function of DAF, the cDNA encoding human DAF was expressed by transfection in Chinese hamster ovary (CHO) cells. Testing of these DAF transfectants in an antibody plus human complement-mediated cytotoxicity assay demonstrated that DAF protects these cells from cytotoxicity, and that the level of protection increases with expression of surface DAF. A cDNA construct encoding a transmembrane version of DAF (DAF-TM) protects CHO transfectants from cytotoxicity with equal efficiency to DAF. This DAF-TM construct used the TM and cytoplasmic domains of membrane cofactor protein (MCP); an alternate TM version of DAF constructed with the TM and cytoplasmic domains of HLA-B44 showed equivalent protection. The protection from cytotoxicity involved a decrease in the deposition of C3 on the cell, consistent with the effect of DAF on the C3 convertase. A second pair of anchor variants, MCP and a GPI-anchored construct, MCP-PI, were also equivalent in their complement protection. The equivalent function of GPI-anchored and TM versions of a protein was not expected based on the hypothesized increased lateral mobility of GPI-anchored proteins, which should confer a functional advantage in contacting ligand, in this case, C3b or C4b, on the cell surface. These data suggest either that GPI-anchored and TM versions of a protein have equal lateral mobility in the membrane, or else that increased lateral mobility is not advantageous to DAF or MCP in carrying out their complement inhibitory roles. Furthermore, DAF and MCP demonstrated approximately equal protection of cells from complement-mediated cytotoxicity, suggesting that DAF and MCP provide overlapping levels of protection to cells against damage mediated by the complement system.
衰变加速因子（DAF）是一种糖基磷脂酰肌醇（GPI）锚定的膜蛋白，通过调节C3转化酶来保护细胞免受补体介导的损伤。为了研究GPI锚在DAF功能中的作用，编码人DAF的cDNA通过转染在中国仓鼠卵巢（CHO）细胞中表达。在抗体加人补体介导的细胞毒性试验中对这些DAF转染子进行检测，结果表明DAF保护这些细胞免受细胞毒性，并且保护水平随着表面DAF的表达而增加。编码跨膜形式DAF（DAF-TM）的cDNA构建体以与DAF相同的效率保护CHO转染子免受细胞毒性。该DAF-TM构建体使用了膜辅因子蛋白（MCP）的跨膜和胞质结构域；用HLA-B44的跨膜和胞质结构域构建的另一种跨膜形式的DAF显示出同等的保护作用。免受细胞毒性的保护作用涉及细胞上C3沉积的减少，这与DAF对C3转化酶的作用一致。另一对锚定变体，MCP和GPI锚定构建体MCP-PI，在补体保护方面也相当。基于推测的GPI锚定蛋白增加的侧向流动性，预计蛋白质的GPI锚定和跨膜形式具有不同的功能，而在这种情况下，GPI锚定蛋白在细胞表面与配体（即C3b或C4b）接触时应具有功能优势。这些数据表明，要么蛋白质的GPI锚定和跨膜形式在膜中具有相等的侧向流动性，要么增加的侧向流动性对DAF或MCP执行其补体抑制作用没有优势。此外，DAF和MCP对细胞免受补体介导的细胞毒性表现出大致相等的保护作用，这表明DAF和MCP为细胞提供了重叠水平的保护，使其免受补体系统介导的损伤。
