Differential response of neuronal cells to a fusion protein of ciliary neurotrophic factor/soluble CNTF-receptor and leukemia inhibitory factor
Differential response of neuronal cells to a fusion protein of ciliary neurotrophic factor/soluble CNTF-receptor and leukemia inhibitory factor
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DOI：
10.1046/j.1432-1033.2002.02977.x
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发表时间：
2002-06-01
期刊：
影响因子：
--
通讯作者：
Rose-John, S
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文献类型：
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作者：
März, P;Özbek, S;Rose-John, S
睫状神经营养因子(CNTF)在体内和体外对运动神经元和神经细胞群具有神经营养活性。在缺乏特定受体α亚基固有表达的靶细胞上，IL-6家族的细胞因子仅在其特定的激动型可溶性受体存在的情况下起作用。在这里，我们报道了CNTF/可溶性CNTF受体(sCNTF-R)融合蛋白(Hyper-CNTF)的构建和表达，该融合蛋白在表达gp130和白血病抑制因子受体(LIF-R)的细胞上具有增强的生物学活性，而不是膜结合的CNTF-R。在基因水平上，人CNTF-R胞外区(氨基酸1-346)的C-末端通过一个甘氨酸残基连接到人CNTF(氨基酸1-186)的N-末端。在COS-7细胞中表达了重组Hyper-CNTF蛋白。Hyper-CNTF能有效诱导稳定表达gp130和LIF-R基因的BAF-3细胞中STAT3的磷酸化和增殖，且呈剂量依赖关系。而在BAF3/gp130/LIF-R细胞上，Hyper-CNTF和LIF具有相似的生物学效应，而在嗜铬细胞瘤细胞(PC12细胞)上，Hyper-CNTF的活性与LIF截然不同。与LIF相反，Hyper-CNTF以时间和剂量依赖的方式刺激PC12细胞突起生长，这与MAP激酶的磷酸化能力有关。这些数据表明，尽管LIF和Hyper-CNTF使用相同的异二聚体受体复合体gp130和LIFR，但只有Hyper-CNTF诱导神经元分化。讨论了高CNTF作为一种超激动型神经营养因子的治疗潜力。
Ciliary neurotrophic factor (CNTF) displays neurotrophic activities on motor neurons and neural cell populations both in vivo and in vitro . On target cells lacking intrinsic expression of specific receptor alpha subunits cytokines of the IL-6 family only act in the presence of their specific agonistic soluble receptors. Here, we report the construction and expression of a CNTF/soluble CNTF-receptor (sCNTF-R) fusion protein (Hyper-CNTF) with enhanced biological activity on cells expressing gp130 and leukemia inhibitory factor receptor (LIF-R), but not membrane-bound CNTF-R. At the cDNA level, the C-terminus of the extracellular domain of human CNTF-R (amino acids 1-346) was linked via a single glycine residue to the N-terminus of human CNTF (amino acids 1-186). Recombinant Hyper-CNTF protein was expressed in COS-7 cells. Hyper-CNTF efficiently induced dose-dependent STAT3 phosphorylation and proliferation of BAF-3 cells stably transfected with gp130 and LIF-R cDNAs. While on BAF3/gp130/LIF-R cells, Hyper-CNTF and LIF exhibited similar biological responses, the activity of Hyper-CNTF on pheochromocytoma cells (PC12 cells) was quite distinct from that of LIF. In contrast to LIF, Hyper-CNTF stimulated neurite outgrowth of PC12 cells in a time- and dose-dependent manner correlating with the ability to phosphorylate MAP kinases. These data indicate that although LIF and Hyper-CNTF use the same heterodimeric receptor complex of gp130 and LIFR, only Hyper-CNTF induces neuronal differentiation. The therapeutic potential of Hyper-CNTF as a superagonistic neurotrophin is discussed.