Mammalian ELAV-like neuronal RNA-binding proteins HuB and HuC promote neuronal development in both the central and the peripheral nervous systems
Mammalian ELAV-like neuronal RNA-binding proteins HuB and HuC promote neuronal development in both the central and the peripheral nervous systems
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DOI：
10.1073/pnas.96.17.9885
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发表时间：
1999-08-17
影响因子：
11.1
通讯作者：
Okano, H
中科院分区：
文献类型：
--
作者：
Akamatsu, W;Okano, HJ;Okano, H
Hu蛋白是哺乳动物胚胎致死性异常视觉系统（ELAV）样神经元RNA结合蛋白，包含3个RNA识别基序，虽然果蝇的ELAV是神经元正确分化和存活所必需的，但Hu基因在哺乳动物神经系统中所起的作用仍然很大程度上是未知的。他们在缺乏神经生长因子的情况下诱导神经元表型，我们已经表征了各种形式的mHuB和mHuC的功能，这些mHuB和mHuC带有点突变或缺失。在第一或第二RNA识别基序（RRMs）的RNP1结构域发生氨基酸交换的mHuC突变体失去了生物活性和RNA结合活性。此外，仅含有第三个RRM的突变体在PC12细胞中未能诱导神经元表型，并且抑制了共转染的野生型mHuB和mHuC的生物活性，从而表现为显性阴性形式。然而，这些突变体不能抑制神经生长因子诱导的PC12细胞分化。此外，我们在E9.5小鼠胚胎中通过电穿孔进入脑管水平，错误表达了野生型和显性阴性的Hu。mHuB和mHuC诱导神经元标志物的异位表达，而mHuB和mHuC的显性阴性形式抑制中枢神经系统运动神经元的分化。从这些结果来看，我们认为Hu蛋白是哺乳动物神经系统神经元分化所必需的。
Hu proteins are mammalian embryonic lethal abnormal visual system (ELAV)-like neuronal RNA-binding proteins that contain three RNA recognition motifs, Although Drosophila ELAV is required for the correct differentiation and survival of neurons, the roles played by the Hu genes in the mammalian nervous system remain largely unknown, To explore the in vivo functions of mouse Hu proteins, we overexpressed them in rat pheochromocytoma PC12 cells, where they induced neuronal phenotype in the absence of nerve growth factor, We have characterized the functions of various forms of mHuB and mHuC bearing point mutations or deletions. Mutants of mHuC that had amino acid exchanges in the RNP1 domain of the first or second RNA recognition motifs (RRMs) lost biologic activity as well as RNA-binding activity. In addition, the mutants containing only the third RRM failed to induce the neuronal phenotype in PC12 cells and inhibited the biologic activity of cotransfected wild-type mHuB and mHuC, thus acting as a dominant-negative form. However, these mutants could not suppress the nerve growth factor-induced differentiation of PC12 cells. Further, we misexpressed wild-type and dominant-negative Hu in E9.5 mouse embryos, by using electroporation into the neural tube at the level of the rhombencephalon. mHuB and mHuC induced the ectopic expression of neuronal markers, whereas the dominant-negative forms of mHuB and mHuC suppressed the differentiation of central nervous system motor neurons, From these results, we suggest that Hu proteins are required for neuronal differentiation in the mammalian nervous system.