Molecular cell-biological studies on differentiation and mitotic arrest of brain neurons

脑神经元分化和有丝分裂停滞的分子细胞生物学研究

基本信息

批准号：
08458253
负责人：
YOSHIKAWA Kazuaki
金额：
$ 4.67万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

Neurons in the vertebrate central nervous system CNS withdraw permanently from the cell cycle immediately after differentiation from their proliferative progenitors. To study the molecular mechanism whereby neurons become postmitotic, we focused on the functional roles and molecular properties of necdin, a neuronal growth suppressor isolated from embryonal carcinoma P19 cells. Results obtained are as follows. [1] Necdin, like retinoblastoma protein (Rb), bound to transforming domains of simian virus 40 (SV40) large T antigen and adenovirus E1A.[2] Necdin bound to E2F-1, a cellular transcription factor that promotes the cell cycle, and suppressed its functions. [3] Ectopic expression of necdin in Rb-dificient SAOS-2 cells suppressed the cell growth, indicating that necdin is a functional substitute for Rb. [4] An antibody against recombinant necdin protein reacted with necdin in the cytoplasm of mouse brain neurons, and the immunoreactivity was the highest in the hypothalamus.[5] The 5'-end sequence of the human necdin gene contains frequent CpG dinucleotides (identified as a CpG island), and methylation in vitro of CpG dinucleotides in the promoter region suppressed the transcriptional activity. [6] The human necdin gene was localized to chromosome 15q11.2-q12, which lies within the region involved in Prader-Willi syndrome, a genomic imprinting-associated neurobehavioral disorder. [7] In summary, necdin suppresses cell growth in a manner similar to that of Rb, and its deficiency may cause the defect of neuronal differentiation.

脊椎动物中枢神经系统 CNS 中的神经元在从其增殖祖细胞分化后立即永久退出细胞周期。为了研究神经元进入有丝分裂后的分子机制，我们重点研究了 necdin（一种从胚胎癌 P19 细胞中分离出来的神经元生长抑制剂）的功能作用和分子特性。得到的结果如下。 [1] Necdin 与视网膜母细胞瘤蛋白 (Rb) 一样，与猿猴病毒 40 (SV40) 大 T 抗原和腺病毒 E1A 的转化结构域结合。 [2] Necdin 与 E2F-1（一种促进细胞周期的细胞转录因子）结合，并抑制其功能。 [3] Rb 缺陷型 SAOS-2 细胞中 necdin 的异位表达抑制了细胞生长，表明 necdin 是 Rb 的功能替代品。 [4] 抗重组necdin蛋白的抗体与小鼠脑神经元细胞质中的necdin发生反应，且在下丘脑中免疫反应性最高。 [5]人necdin基因的5'端序列含有频繁的CpG二核苷酸（鉴定为CpG岛），启动子区CpG二核苷酸的体外甲基化抑制了转录活性。 [6] 人类 necdin 基因定位于染色体 15q11.2-q12，该区域位于与 Prader-Willi 综合征（一种基因组印记相关的神经行为障碍）有关的区域。 [7]综上所述，necdin以与Rb类似的方式抑制细胞生长，其缺乏可能导致神经元分化缺陷。