Overexpression of HuD accelerates neurite outgrowth and increases GAP-43 mRNA expression in cortical neurons and retinoic acid-induced embryonic stem cells in vitro
Overexpression of HuD accelerates neurite outgrowth and increases GAP-43 mRNA expression in cortical neurons and retinoic acid-induced embryonic stem cells in vitro
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DOI：
10.1006/exnr.2000.7599
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发表时间：
2001-04-01
影响因子：
5.3
通讯作者：
Perrone-Bizzozero, NI
中科院分区：
文献类型：
--
作者：
Anderson, KD;Sengupta, J;Perrone-Bizzozero, NI
神经元特异性RNA结合蛋白HuD与GrAP-43 mRNA 3 '非翻译区（3 ' UTR）的富含U的调节元件结合并稳定mRNA，我们之前已经证明PC 12细胞中HuD的过表达增加了GAP-43蛋白表达并诱导自发形成多个神经突（K，D，安德森等人，2000，神经化学杂志，75：1103-1114）。在这项研究中，我们研究了HuD过度表达的影响，神经突起生长的初始阶段和GAP-43基因表达使用两个体外系统：E19大鼠皮层神经元和维甲酸（RA）诱导的胚胎干细胞（ES）。体外皮质神经元的正常神经突生长发生在3天的时间内，伴随着GAP-43和HuD表达的增加。皮质细胞用含有正义方向HuD cDNA的复制缺陷型HSV-1载体（HSV-HuD）感染。免疫细胞化学分析显示，过量HuD处理24 h后，GAP-43阳性细胞数增加3倍，并发生形态学分化。使用原位杂交，我们发现HuD表达增加导致GAP-43 mRNA水平增加两倍。类似地，发现在RA诱导的胚胎干细胞中HuD过表达增加了GAP-43阳性细胞的数量。总之，我们的研究结果表明，HuD的功能，在神经突生长的方式，由于启动，至少部分，其调节GAP-43的表达。(C)北京：科学出版社.
The neuron-specific RNA-binding protein HuD binds to a U-rich regulatory element of the 3 ' untranslated region (3 ' UTR) of the GrAP-43 mRNA and stabilizes the mRNA, We have previously shown that overexpression of HuD in PC12 cells increases GAP-43 protein expression and induces the spontaneous formation of multiple neurites (K, D, Anderson et al, 2000, J, Neurochem, 75: 1103-1114). In this study, we examined the effects of HuD overexpression on the initial stages of neurite outgrowth and on GAP-43 gene expression using two in vitro systems: E19 rat cortical neurons and retinoic acid (RA)-induced embryonic stem (ES) cells. Normal neurite outgrowth of cortical neurons in vitro occurs over a 3-day period with a concomitant increase in GAP-43 and HuD expression. Cortical cells were infected with a replication-deficient HSV-1 vector containing the HuD cDNA in the sense orientation (HSV-HuD). Overexpression of HuD accelerated the formation of neurites, Immunocytochemical analysis showed that excess HuD resulted in a threefold increase in the number of GAP-43-positive cells undergoing morphological differentiation after 24 h of treatment. Using in situ hybridization, we found that the increased HuD expression resulted in a twofold increase in the levels of GAP-43 mRNA Similarly, overexpression of HuD in RA-induced embryonic stem cells was found to increase the number of GAP-43-positive cells undergoing process outgrowth. In conclusion, our results demonstrate that HuD functions in the initiation of neurite outgrowth in a manner due, at least in part, to its regulation of GAP-43 expression. (C) 2001 Academic Press.