权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Functional analysis of BTEB and BTEB2 transcription factors by gene-targeting technology

基因打靶技术对BTEB和BTEB2转录因子的功能分析

基本信息

批准号：
13480200
负责人：
FUJII Yoshiaki
金额：
$ 7.94万
依托单位：
University of Tsukuba (2002)Tohoku University (2001)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13480200/
关键词：
BTEB BTEB2 targeting vectors FGF4 GCbox consensus sequence gene-knockout mice cerebellum hippocampus ターゲッティングペクターホモ欠失マウス in vitro 標的遺伝子 β-gal T_3

项目摘要

Mouse BTEB and BTEB2 are transcription factors with three-time-repeated C_2H_2 zinc finger motif and bind the GC-box consensus sequence. In order to investigate their physiological functions, we have constructed targeting vectors by replacing the first exon with structural genes for β-gal in translational coding phase and neo and made BTEB- and BTEB2-deficient mice by homologous recombination technology using these targeting vectors. Concerning the BTEB-null mice, they, either male or female, were born apparently normal according to the Mendelian genetics from mating between heterozygous BTEB (+/-) dams and grew normal. The male and female offspring were fertile. Judging from the expression pattern of BTEB during the mouse development showing that the dramatically increased expression of BTEB was observed in Purkinye cells of the cerebellum and pyramidal cell layers of the hippocampus at P7 when synapses start to form in the brain, we investigated the memory and motor activity. Although general behavioral activities such as locomotion, rearing, and movement were not so much affected in the BTEB (-/-) mutant mice, they showed clearly reduced activity levels in rotorod and contextual fear conditioning tests, probably due to defective functions of the cerebellum, and hippocampus, respectively. On the other hand, homozygous BTEB-2 (-/-) mice were not born, while heterozygous BTEB2 (+/-) mice were born normally. Detailed analysis of embryos revealed that fertilized BTEB (-/-) oocytes developed apparently normally until E3.5 and died at E4.5. One of the causes for the early embryonic death was considered to be defective expression of FGF4 which is known to be important factor for the early embryonic development and was clarified to be a target gene of BTEB2.

小鼠BTEB和BTEB2是具有三次重复的C_2H_2锌指基序的转录因子，并与GC-box共同序列结合。为了研究它们的生理功能，我们构建了靶向载体，将β-Gal的第一个外显子替换为翻译编码阶段和NEO的结构基因，并利用这些靶向载体通过同源重组技术建立了BteB和BteB_2缺陷小鼠。对于BTEB基因缺失的小鼠，根据孟德尔遗传学，它们出生时明显正常，无论是雄性还是雌性，来自杂合子BTEB(+/-)母鼠之间的交配，并生长正常。雄性和雌性后代都能生育。根据BTEB在小鼠发育过程中的表达模式显示，当突触开始在大脑中形成时，BTEB在P7的小脑Purkinye细胞和海马锥体细胞层中的表达显著增加，我们研究了BTEB在记忆和运动活动中的作用。虽然BTEB(-/-)突变小鼠的一般行为活动，如运动、站立和运动没有受到太大影响，但它们在旋转和情景恐惧条件反射测试中显示出明显的活动水平降低，可能分别是由于小脑和海马体的功能缺陷。另一方面，纯合子BTEB-2(-/-)小鼠没有出生，而杂合子BTEB2(+/-)小鼠正常出生。对胚胎的详细分析表明，受精的BTEB(-/-)卵母细胞在E3.5之前发育明显正常，并在E4.5死亡。早期胚胎死亡的原因之一是FGF4的表达缺陷，FGF4是胚胎早期发育的重要因素，并被阐明为BTEB2的靶基因。