Creation of CTIP1 and CTIP2 Null Mice

本提案的目标（单期，12个月）是建立不表达COUP tf相互作用蛋白1和2（分别为CTIP1和CTIP2）的小鼠，目的是阐明每种蛋白在胚胎发生和成年动物中的作用。这个休假期也将为PI提供一个机会，发展与小鼠基因组操作相关的技术专业知识，包括胚胎干细胞的同源重组和培养，囊胚注射和所得小鼠的基因分型/分析。该项目的目标是在俄勒冈州立大学校园内建立这种技术，俄勒冈州立大学目前缺乏，但非常需要一种制造基因敲除和转基因动物的设施。CTIP1和CTIP2是C2H2锌指蛋白新家族的两个成员，它们是在酵母双杂交筛选中分离出来的，该筛选旨在鉴定能够与孤儿核受体COUP-TF家族成员相互作用的蛋白。CTIP1和CTIP2在大脑中都有高表达，COUP-TF蛋白家族的所有成员也是如此。研究人员对CTIP1进行了详细的研究，发现CTIP1具有自主的转录调控结构域，并在哺乳动物细胞中以一种对组蛋白去乙酰化酶抑制剂trichostatin a不敏感的方式增强ARP1 （COUP-TF家族成员）的转录抑制活性。这一发现表明，ARP1介导了转录抑制。哺乳动物细胞中的CTIP1复合物可能在机制上不同于其他核受体。本文所描述的工作将有助于阐明CTIP蛋白在生物环境中的功能，并定义COUP-TF家族成员通过C2H2锌指蛋白新家族在胚胎发生和成年动物中发挥转录调节活性的潜在分子机制。

The objective of this proposal (single term, 12 months) is to create mice null for expression of COUP TF-Interacting Proteins 1 and 2 (CTIP1 and CTIP2, respectively) toward the goal of elucidating the role of each protein during embryogenesis and in the adult animal. This sabbatical period will also afford an opportunity for the PI to develop expertise in the techniques associated with genomic manipulations in the mouse including homologous recombination in and culture of embryonic stem cells, blastocyst injection and genotyping/analysis of the resultant mice. The PI's goal is to establish such technology on the campus of Oregon State University, which is currently devoid, but in great need, of a facility for creation of knock-out and transgenic animals. CTIP1 and CTIP2 represent two members of a novel family of C2H2 zinc finger proteins that were isolated in a yeast two-hybrid screen conducted to identify proteins capable of interaction with members of the COUP-TF family of orphan nuclear receptors. Both CTIP1 and CTIP2 are highly expressed in brain, as are all members of the COUP-TF family of proteins. CTIP1 has been examined in detail and was found to harbor autonomous transcriptional regulatory domains and to potentiate the transcriptional repression activity of ARP1, a COUP-TF family member, in mammalian cells in a manner that was not sensitive to reversal by the histone deacetylase inhibitor, trichostatin A. This finding suggests that transcriptional repression mediated by ARP1.CTIP1 complexes in mammalian cells may be mechanistically distinct from that of other nuclear receptors. The work described herein will facilitate elucidation of the function of the CTIP proteins in an organismal context as well as definition of the underlying molecular mechanisms by which COUP-TF family members, acting through this novel family Of C2H2 zinc finger proteins, may exert transcriptional regulatory activity during embryogenesis and in the adult animal.