PDX-1 IN MAMMALIAN PANCREATIC DEVELOPMENT

我们的长期目标是提供与多潜能胰腺干细胞的鉴定、纯化、扩增和分化相关的信息，这可能会产生适合胰腺替代疗法的材料，并增加我们对II型糖尿病细胞缺陷的理解，并最终帮助抵消。尽管这些领域的一些进展最近来自对具有编码胰腺中表达的转录因子的失活基因的小鼠的研究，但我们对胚胎发生过程中内分泌/外分泌分化以及成人细胞类型和功能维持的机制理解仍然处于初级阶段。我们提出了一组有针对性的实验，旨在填补其中一些空白。首先，我们将对胰腺必需基因 pdx-1 进行精确的功能分析。全局 pdx-1 失活会阻碍胰腺发育，我们现在将确定 pdx-1 在胰腺细胞分化后期、外分泌、β 和非 β 细胞基因中的作用，表明 pdx-1 转录调控是控制 β 细胞功能的主要焦点。我们已经鉴定出可能的 pdx-1 顺式调控元件，其中包含转录因子（包括 HNF3beta 和最近描述的 MODY 易感因子）的结合位点。我们将确定这些保守的顺式调控元件在体内的作用。第三，我们将开发遗传谱系追踪技术来表征内分泌细胞个体发育。具体目标是：(1)确定灭活pdx-1对选定的内分泌和外分泌细胞的影响。 (2)确定pdx-1中保守顺式调控序列的体内作用，开始剖析控制胰腺形成和功能的转录调控网络。 (3) 开发遗传谱系追踪技术，以严格了解胰腺细胞个体发育，特别是内分泌途径。

Our long-term goals are to produce information relevant to the identification, purification, expansion and differentiation of multi- potential pancreatic stem cells, which may lead to material suitable for pancreas replacement therapy, and increase our understanding of, and eventually help to offset, the cellular defects in type II diabetes. Although some progress in these areas has come recently from studies of mice with inactivated genes that encode transcription factors expressed in the pancreas, our mechanistic understanding of endocrine/exocrine differentiation during embryogenesis, and maintenance of cell type and function in adults, is still rudimentary. We propose a focused set of experiments directed at filling some of these gaps. First, we will carry out a precise functional analysis of an essential pancreatic gene, pdx-1. Global pdx-1 inactivation blocks pancreas development, and we will now determine the role of pdx-1 in later stages of pancreatic cell differentiation, in the exocrine, beta and non-beta cell genes, indicating that pdx-1 transcriptional regulation is a major focus for the control of beta cell function. We have identified likely pdx-1 cis-regulatory elements that contain binding sites for transcription factors that include HNF3beta and the recently described MODY susceptibility factors. We will determine the role of these conserved cis-regulatory elements in vivo. Third, we will develop genetic lineage tracing techniques to characterized endocrine cell ontogeny. Specific aims are: (1) to determine the effect of inactivating pdx-1 in selected endocrine and exocrine cells. (2) to determine in vivo the role of conserved cis-regulatory sequences in pdx-1, to begin to dissect the transcriptional regulatory networks controlling pancreas formation and function. (3) Develop genetic lineage tracing techniques for a rigorous understanding of pancreatic cell ontogeny, with special reference to the endocrine pathway.