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Glial cell line-derived neurtftrophic factor (GDNF) inducible genes and neuronal differentiation

胶质细胞源性神经营养因子（GDNF）诱导基因和神经元分化

基本信息

批准号：
12670199
负责人：
ICHIHARA Masatoshi
金额：
$ 2.18万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670199/
关键词：
RET GDNF neurite o utgrowth neuronal differentiation inducible gene zinc fingerprotein

项目摘要

Glial cell line-derived neurotrophic factor (GDNF) family ligands promote the growth and survival of several population of CMS and PNS neuron via activation of the RET tyrosine kinase. We searched for immediate-early GDNF responsive genes by mRNA differential display technique using TGW neuroblastoma cells and have isolated fourteen GDNF inducible genes including c-fos, CREB and CREM. We focused on two novel genes. One novel gene was induced in the fast kinetics manner like c-fos and its protein sequence deduced from CDNA ORF contains zinc finger repeat in the C-terminal moiety and KRAB domain, BTB/POZ domain in N-terminal, respectively. In northern blot of human multiple tissues, its expression was confirmed in adult kidney as well as skeletal muscle and heart. Interestingly, immunohistochemical staining of mouse embryo showed that ureteric bud epithelia were stained similarly to RET expression but not in adult kidney. We are now evaluating its function using antisense oligonucleotide in the organ culture of the mouse embryonic kidneys. These results suggest that this novel zinc finger protein may play a role in nephrogenesis under Ret/GDNF system in addition to neuronal cells. We also focused on the other unknown gene. Molecular cloning of the full length CDNA of this gene revealed that it was the human homologue which counterpart of budding yeast had an important role in the many cellular process including miosis, RNA metabolism and telomere function. Northern and western blot showed that this gene was also highly induced in the late-phase in the GDNF stimulation. Immunohistochemical staining showed that this gene was expressed in extending neurite in neuroblastoma cells and in neuronal tissues of adult mice and mouse embryo in the later term of embryogenesis. We also found its co-localization with tubulin, suggesting its important role in neurite outgrowth through the mechanism like tubulin stabilization.

胶质细胞源性神经营养因子（GDNF）家族配体通过激活RET酪氨酸激酶促进几种CMS和PNS神经元群体的生长和存活。本研究利用mRNA差异显示技术，以TGW神经母细胞瘤细胞为研究对象，筛选出14个GDNF诱导基因，包括c-fos、CREB和CREM。我们专注于两个新基因。其中一个新基因是以类似于c-fos的快动力学方式诱导产生的，其蛋白质序列在C端含有锌指重复序列，在N端分别含有KRAB结构域和BTB/POZ结构域。在人多种组织的北方印迹中，证实了其在成人肾脏以及骨骼肌和心脏中的表达。有趣的是，小鼠胚胎的免疫组织化学染色显示输尿管芽上皮染色类似于RET表达，但在成年肾脏中没有。我们正在利用反义寡核苷酸在小鼠胚胎肾脏器官培养中评价其功能。这些结果表明，这种新的锌指蛋白可能在Ret/GDNF系统下的肾发生中发挥作用，除了神经细胞。我们还关注了另一个未知基因。该基因全长cDNA的克隆表明，该基因是人类同源基因，与芽殖酵母类似，在减数分裂、RNA代谢和端粒功能等细胞过程中发挥重要作用。北方和西方印迹表明，该基因在GDNF刺激的后期也被高度诱导。免疫组织化学染色显示，该基因在神经母细胞瘤细胞的延伸神经突中以及成年小鼠和胚胎发育后期小鼠胚胎的神经元组织中均有表达。我们还发现了它与微管蛋白的共定位，这表明它通过微管蛋白稳定机制在神经突生长中起重要作用。