Endocrine cell induction during pancreas

在胰腺发育过程中，祖细胞形成朗格汉斯岛需要Delta/Notch信号通路的协同激活。尽管这一途径与维持胰腺祖细胞的多能性和支持细胞命运决定有关，但人们对其了解甚少。为了更好地理解胰腺发育过程中发生的这些和其他事件，能够轻松检查发育中的小鼠胰腺中功能获得和功能丧失突变的影响是很重要的。在可能的情况下，将这些突变有条件地激活或不激活也是有用的。因此，该项目的重点是在小鼠胚胎干细胞中使用和进一步开发磁带交换技术。这种策略加速了胚胎干细胞中高度定义的基因突变的产生，然后这些突变可以用来制造新的小鼠，或者作为学习如何指导胚胎干细胞向胰腺细胞或β细胞分化的工具。即将开发的试剂，以及即将进行的实验，有望加速对胰腺内产生内分泌细胞的分子事件的理解。在Aim 1中，我们将利用Ptf1a基因位点控制下表达黄色荧光蛋白的小鼠来研究胰腺祖细胞的生长和分化能力。对于目标2，我们将探索

The formation of Islets of Langerhans from progenitor cells during pancreas development requires the coordinated activation of the Delta/Notch signaling pathway. This pathway, although it has been implicated in both maintaining multipotency and supporting cell fate decisions of pancreatic progenitor cells, is poorly understood. In order to better understand these and other events that occur during pancreas development, it is important to be able to easily examine the effects, within the developing mouse pancreas, of both gain-of-function and loss-of-function mutations. When possible, it is also useful for these mutations to be conditionally active or inactive. Thus, this project is focused on both the use and further development of cassette exchange technologies in mouse ES cells. This strategy accelerates the production of highly defined gene mutations in ES cells, which can then be used either to make new mice or as tools for learning how to direct the differentiation of ES cells towards a pancreatic or beta cell fate. The reagents that will be developed, as well as the experiments that will be performed, promise to accelerate progress towards an understanding of the molecular events that give rise to endocrine cells within the pancreas. In Aim 1 we will explore the growth and differentiation capacity of pancreatic progenitor cells using mice in which yellow fluorescent protein is expressed under control of the Ptf1a gene locus. For Aim 2 we will explore alterations in Delta/Notch signaling by expressing three different fringe genes under control of the Ptf1a gene locus. In Aim 3 we will assess the effects of a global knock-out of Insm1 while also generating a new loxed cassette acceptor allele using an improved strategy. For Aim 4 we will generate and test new mouse ES cell lines that will facilitate learning how to direct the differentiation of these cells towards a pancreatic beta cell fate. Lastly, in Aim 5 we will establish a new cassette exchange system that makes use of phiC31 integrase to allow rapid insertion of genes into the ROSA26 locus and their subsequent induction in a site- and temporal-specific manner by Cre.