Embryonic dvelopment and the orphan receptor GCNF

我们克隆了一个新的孤儿核受体w3，其胚胎表达模式表明它参与调节重要的发育过程。该项目的长期目标是了解孤儿核受体GCNF在胚胎发育中的生理作用。为此，我们创建了一个小鼠模型，其中GCNF功能的完全丧失导致妊娠中期胚胎死亡，对许多发育过程和系统产生深远影响。这包括主要的模式缺陷，包括前后轴断裂，导致后脑尾侧严重截断，躯体发育停止，中胚层分化缺陷，以及后腹侧器官发生失败。我们假设GCNF是一种重要的发育受体和转录因子，通过控制发育重要基因的表达来调节胚胎轴的形成和器官发生。为了验证这一假设，我们将利用GCNF敲除小鼠模型来解剖这种孤儿受体在胚胎发育中的作用。为实现这一目标，我们提出了三个具体目标。1)完成GCNF基因敲除胚胎缺陷表型的表征。LacZ报告基因将被敲入GCNF位点，以帮助表征GCNF敲除表型。完成前-后轴型缺损、器官发生异常和中胚层分化缺损的表征。2)随后，我们将确定GCNF在三个胚层中是否以细胞自主或非自主的方式作用以产生观察到的表型。使用LacZ敲入小鼠，将产生含有标记有β - gal的GCNF (-/-) ES细胞和野生型ES细胞的聚集嵌合体来回答这个问题。3)我们建议鉴定GCNF在胚胎发生中重要的靶基因。我们将确定GCNF缺失对适当突变型和野生型胚胎表达的影响。这些实验的完成将使我们更好地了解GCNF通过控制基因表达在调节胚胎发育中的作用。

We have cloned a novel orphan nuclear receptor w3hose embryonic expression pattern suggests that it is involved in regulating important developmental processes. The long term goal of this project is to understand the physiological role of the orphan nuclear receptor GCNF in embryonic development. To this end we have created a mouse model in which complete loss of GCNF function leads to embryonic lethality mid- gestation, with profound effects on many developmental processes and systems. This includes a major patterning defect involving disruption of the anterior-posterior axis causing a severe truncation caudal to the hindbrain, a halt in somitogenesis and defective differentiation of mesoderm, and failure of posterior ventral organogenesis. We hypothesize that GCNF is a developmentally important receptor and transcription factor that regulates embryonic axis formation and organogenesis by controlling the expression of developmentally important genes. To test this hypothesis we will utilize the GCNF knock- out mouse model to dissect the role of this orphan receptor in embryonic development. To achieve this goal we propose three specific aims. 1) Complete the characterization of the defective phenotype of the GCNF knockout-out embryos. The LacZ reporter gene will be knocked into the GCNF locus to aid in the characterization of the GCNF knock-out phenotype. The characterization of the patterning defects anterior- posterior axis, organogenesis abnormalities and the mesodermal differentiation defects will also be completed. 2) Subsequently, we will determine whether GCNF is acting in a cell autonomous or non- autonomous manner in each of the three germ layers to generate the observed phenotype. Using the LacZ knock-in mice, aggregation chimeras, containing GCNF (-/-) ES cells tagged with beta-Gal and wild type ES cells will be generated to answer this question. 3) We propose to identify GCNF target genes important for embryogenesis. We will determine effects of the loss of GCNF on the expression of appropriate mutant and wild type embryos. The completion of these experiments will yield greater understanding of the role GCNF plays in regulating embryonic development by controlling gene expression.