EXPRESSION/FUNCTION OF C FOS AND C JUN IN DEVELOPMENT

C FOS 和 C JUN 在发育中的表达/功能

• 批准号: 3328945
• 负责人: VIRGINIA E. PAPAIOANNOU
• 金额: $ 26.88万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-09-01 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3328945
• 关键词: congenital disorders; developmental genetics; embryo implantation; gametogenesis; gene expression; gene mutation; genetic manipulation; genetic recombination; genetically modified animals; heterozygote; immunocytochemistry; in situ hybridization; laboratory mouse; molecular cloning; protooncogene; transposon /insertion element

The goals of this proposal are to investigate the role of proto-oncogenes in normal mammalian development and to understand how aberrations in the expression of these genes could result in pregnancy wastage or congenital malformations. C-fos and c-jun proto-oncogenes function together to regulate gene expression at the molecular level and have been chosen for this study as proto-oncogenes implicated in development. C-fos is expressed in a tissue-specific manner during embryogenesis and shows high levels of expression in extraembryonic tissues. These tissues play an essential role in implantation and in the survival of the embryo during the postimplantation period. It is our hypothesis that tissue-specific expression reflects a functional requirement for the c-fos gene product. We further postulate that c-jun will be expressed in a similar manner and will also be required during development. The specific aims of this proposal are to clarify the extent of c-fos and c-jun expression throughout the pre- and early postimplantation period in the mouse and examine the functional role of these genes in development. This will be accomplished by the following specific aims: Specific Aim I. We will detail the expression of c-fos and c-jun during pre- and postimplantation mouse development using in situ hybridization for RNA localization and immunohistochemistry for localization of the proteins. Specific Aim 2. We will produce embryonic stem (ES) cell lines heterozygous for mutated c-fos and c-jun by gene targeting via homologous recombination. Preliminary results have been successful in targeting c-fos and one mutated clone is now available. Specific Aim 3. We will then produce transgenic animals heterozygous and homozygous for mutated C-fos or c-jun by way of ES cell chimeras, in order to determine the effect of a lower dose or absence of c-fos or c-jun gene product on development. One series of chimeras is already being test-bred to produce heterozygous mice. If we determine that heterozygous cells are not capable of completing gametogenesis in a chimera, an alternative, rescue protocol will be used that will allow the study of gametogenesis in heterozygous males.

这项计划的目标是研究原癌基因的作用 在正常哺乳动物的发育过程中， 这些基因的表达可能导致妊娠浪费或先天性 畸形C-fos和c-jun原癌基因共同发挥作用， 在分子水平上调节基因表达， 这项研究认为原癌基因与发育有关。 C-fos是 在胚胎发生过程中以组织特异性方式表达， 在胚外组织中的表达水平。这些组织发挥着 在植入和胚胎存活过程中的重要作用 植入后时期我们假设组织特异性 表达反映了对c-fos基因产物的功能需求。我们 进一步假设c-jun将以类似的方式表达，并且将 在发展过程中也需要。 本提案的具体目的是澄清c-fos的范围， c-jun在整个植入前和植入后早期的表达 研究这些基因在发育中的功能作用。 这将通过以下具体目标来实现： 具体目标一。 我们将详细介绍c-fos和c-jun在预处理和 RNA原位杂交技术在小鼠胚胎着床后发育中的应用 定位和免疫组织化学用于蛋白质的定位。 具体目标2。 我们将生产胚胎干细胞系 通过同源基因靶向，对突变的c-fos和c-jun进行杂合 重组初步结果已经成功地针对c-fos 现在有一个变异的克隆体 具体目标3。然后我们将 产生对于突变的C-fos杂合和纯合的转基因动物，或 c-jun通过ES细胞嵌合体的方式，以确定 低剂量或缺乏c-fos或c-jun基因产物对发育的影响。一 一系列的嵌合体已经被用来培育杂合子小鼠。 如果我们确定杂合细胞不能完成 嵌合体中的配子发生，将使用替代的拯救方案 这将允许研究杂合子男性的配子发生。