PDX-1 REGULATION OF PANCREAS DEVELOPMENT/DIFFERENTIATION

1型和2型糖尿病的标志是胰腺β细胞群不能产生足够的胰岛素。 胰岛素以满足代谢需求。PDX-1（Pancreas Duodenal homeoboX-1）是一个同源结构域 一种在控制β-淀粉样蛋白发育的转录层次中处于枢轴位置的转录因子 细胞团而PDX-1的纯合突变导致小鼠和人类的胰腺发育不全， 杂合PDX-1基因突变损害β细胞功能和存活，导致葡萄糖耐受不良， 在小鼠和人类中的糖尿病（早期[MODY 4]-和晚发型2型糖尿病）。PDX-1单倍不足 在几种胰岛素抵抗小鼠模型中显著损害胰岛代偿反应， 受损的胰岛素分泌和/或β细胞量扩增和β细胞存活。还有很多东西要学 关于PDX-1调节基因转录的分子机制以及相关的PDX-1 介导其对β细胞的关键作用的靶点。我们假设PDX-1的C端， 在家族性人2型糖尿病中突变的，是胎儿晚期/早期正常BEA细胞扩增所必需的。 新生儿期此外，我们假设在成人β细胞中鉴定PDX-1靶点将使 2型糖尿病患者胰岛代偿机制的研究进展这些假设将是 目的1：确定PDX-1 C-末端的体内和分子作用：我们将 表征表达缺乏C末端的过早截短的PDX-1的PDX-1 Cterm-/-小鼠 通过评估谱系特化，β细胞增殖和存活，β细胞身份的关键标志物， 和胰岛素分泌。生物化学实验将解决C-末端在蛋白质周转中的作用， 亚细胞定位和靶启动子占有率。C-末端的点突变和快速的 将对斑马鱼的选定突变进行生物筛选。目的2：鉴定PDX-1的转录 靶点是成人β细胞我们将描述一种位于基因上游的新型PDX-1增强子， PDX-1与出生后功能高度相关，我们将探讨其在介导β细胞营养因子中的作用。 GLP-1和胰岛补偿胰岛素抵抗的作用。此外，我们将执行“芯片上的ChIP” 使用成年小鼠胰岛染色质进行分析。一个互补的生物信息学方法将重点 对PDX-1增殖相关靶基因的亚组的影响，这些靶基因由PDX-1：PBX异二聚体调节。 总之，拟议的研究将解决有关PDX-1在β细胞中功能的重要问题。 生物学，特别是其在胰岛素抵抗的β细胞补偿中的作用。

A hallmark of both type 1 and type 2 diabetes is the failure of pancreatic beta cell mass to produce sufficient insulin to meet metabolic demands. PDX-1 (Pancreas Duodenal homeoboX-1) is a homeodomain transcription factor that is pivotally positioned in the transcriptional hierarchy governing the development of beta cell mass. Whereas homozygous mutation of PDX-1 causes pancreatic agenesis in both mice and humans, heterozygous PDX-1 gene mutations impair beta-cell function and survival, leading to glucose intolerance and diabetes in mice and in humans (early [MODY4]- and late-onset type 2 diabetes). PDX-1 haploinsufficiency markedly impairs the islet compensatory response in several insulin resistance mouse models, due to impaired insulin secretion and/or beta cell mass expansion and beta cell survival. Much remains to be learned about the molecular mechanisms whereby PDX-1 regulates gene transcription and about the relevant PDX-1 targets that mediate its critical actions on the beta cell. We hypothesize that the PDX-1 C terminus, which is mutated in familial human type 2 diabetes, is required for normal bea cell expansion in the late fetal/early neonatal period. Further, we hypothesize that the identification of PDX-1 targets in the adult beta cell will give insight into mechanisms of islet compensation in patients with type 2 diabetes. These hypotheses will be tested as follows: Aim 1: Determine the in vivo and molecular role of the PDX-1 C-terminus: We will characterize PDX-1 Cterm-/- mice that express a prematurely truncated PDX-1 lacking the C-terminal domain by assessing lineage specification, beta cell proliferation and survival, critical markers of beta cell identity, and insulin secretion. Biochhemical experiments will address the role of the C-terminus in protein turnover, subcellular localization and target promoter occupancy. Point mutagenesis of the C-terminus and a rapid biological screen of selected mutations in zebrafish will be carried out. Aim 2: Identify PDX-1 transcriptional targets in the adult beta cell. We will characterize a novel PDX-1 enhancer located upstream of a gene with high relevance to the postnatal functions of PDX-1, and we will explore its role in mediating the beta cell trophic effects of GLP-1 and in islet compensation for insulin resistance. Further, we will perform a "ChIP on chip" analysis using adult mouse pancreatic islet chromatin. A complementary bioinformatics approach will focus on a subset of PDX-1 proliferation related target genes that are regulated by PDX-1: PBX heterodimers. Together, the proposed studies will address important questions about the functions of PDX-1 in beta cell biology, with particular relevance to its role in beta cell compensation for insulin resistance.