Heinrich J N, Bravo R
Department of Molecular Biology, Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, New Jersey 08543-4000, USA.
J Biol Chem. 1995 Nov 24;270(47):28014-7. doi: 10.1074/jbc.270.47.28014.
We have demonstrated that the mouse chemokine N51, also known as KC, can compete for 125I-human interleukin-8 (IL-8) binding to NIH 3T3 cells expressing the human IL-8 receptor beta (NIH-IL-8R beta) but not the IL-8 receptor alpha (NIH-IL-8R alpha). In addition, we used the chimeras between N51 and IL-8 described previously (Heinrich, J. N., O'Rourke, E. C., Chen, L., Gray, H., Dorfman, K. S., and Bravo, R. (1994) Mol. Cell. Biol. 14, 2849-2861; Heinrich, J. N., and Bravo, R. (1995) J. Biol. Chem. 270, 4987-4989) to evaluate possible contributions of equivalent domains from each chemokine to binding and specificity. Specifically, the amino acid sequences between cysteines 2 and 3 or between cysteines 3 and 4 or the alpha-helical C-terminal end (domains I, II, and III, respectively) of one of the chemokines was exchanged with the corresponding sequence of the other and vice versa. Chimeras of IL-8 containing either domain II or III of N51 behaved similarly, but not identically, to IL-8 in competing 125I-IL-8 binding with both NIH-IL-8R alpha cells and NIH-IL-8R beta cells. The IL-8 chimera containing domain I of N51 did not compete. On the other hand, N51 competes 125I-IL-8 binding with NIH-IL-8R beta but not NIH-IL-8R alpha cells. The N51 chimera containing domain I of IL-8 was an agonist with NIH-IL-8R alpha cells and was an even more potent agonist with NIH-IL-8R beta cells. On the latter cells it was more potent than either IL-8 or N51. The N51 chimera containing domain II of IL-8, compared with N51, was a partial agonist with NIH-IL-8R alpha cells but was equivalent to N51 with NIH-IL-8R beta cells. However, N51 chimera containing domain III of IL-8 was a partial agonist with both cells. The results are consistent with the observations we originally made with human neutrophils and the NIH mouse IL-8R beta cells, i.e. domain I confers binding specificity for IL-8 and domains II and III of IL-8 and N51 may be interchangeable but they are not equivalent. Although we originally hypothesize that domains II and III confer binding specificity to N51, these results emphasize the role of domain III.
我们已经证明，小鼠趋化因子N51（也称为KC）可以竞争125I-人白细胞介素8（IL-8）与表达人IL-8受体β（NIH-IL-8Rβ）而非IL-8受体α（NIH-IL-8Rα）的NIH 3T3细胞的结合。此外，我们使用了先前描述的N51和IL-8之间的嵌合体（Heinrich, J. N., O'Rourke, E. C., Chen, L., Gray, H., Dorfman, K. S., and Bravo, R. (1994) Mol. Cell. Biol. 14, 2849 - 2861; Heinrich, J. N., and Bravo, R. (1995) J. Biol. Chem. 270, 4987 - 4989）来评估每种趋化因子的等效结构域对结合和特异性的可能贡献。具体而言，将其中一种趋化因子的半胱氨酸2和3之间、半胱氨酸３和4之间的氨基酸序列或α-螺旋C末端（分别为结构域I、II和III）与另一种趋化因子的相应序列进行交换，反之亦然。含有N51结构域II或III的IL-8嵌合体在与NIH-IL-8Rα细胞和NIH-IL-8Rβ细胞竞争125I-IL-8结合时，其行为与IL-8相似，但不完全相同。含有N51结构域I 的IL-8嵌合体不参与竞争。另一方面，N51与NIH-IL-8Rβ细胞竞争125I-IL-8结合，但不与NIH-IL-8Rα细胞竞争。含有IL-8结构域I 的N51嵌合体对NIH-IL-8Rα细胞是一种激动剂，对NIH-IL-8Rβ细胞是一种更强效的激动剂。在后者细胞上，它比IL-8或N51都更有效。与N51相比，含有IL-8结构域II的N51嵌合体对NIH-IL-8Rα细胞是一种部分激动剂，但对NIH-IL-8Rβ细胞与N51相当。然而，含有IL-8结构域III的N51嵌合体对两种细胞都是一种部分激动剂。这些结果与我们最初用人中性粒细胞和NIH小鼠IL-8Rβ细胞所做的观察结果一致，即结构域I赋予对IL-8的结合特异性，IL-8和N51的结构域II和III可能是可互换的，但它们并不等同。尽管我们最初假设结构域II和III赋予N51结合特异性，但这些结果强调了结构域III的作用。
