Intestinal secretory lineage development and function

描述（由申请人提供）： 在肠上皮中，Lieberkuhn隐窝内的增殖和凋亡之间的平衡维持绒毛高度和隐窝深度;这种平衡的破坏导致癌症。在隐窝内，干细胞产生吸收细胞和分泌细胞。几个基因已经牵连在这个过程中，但描绘了一个有序的层次结构控制肠上皮分化是缺乏的。这个层次中的一个基因Math 1是分泌谱系发育所必需的;缺乏Math 1的小鼠产生吸收性肠上皮细胞，但不产生分泌谱系。该提案将测试的假设，数学1依赖的分泌谱系所需的局部增殖稳态在肠道中，和分泌谱系分化的遗传途径的划定将澄清肠道稳态的机制。首先，我将通过基于微阵列的Math 1-/-隐窝和Gfi 1无效隐窝的表达谱来表征肠分化的遗传途径，Gfi 1是一种假定的Math 1靶基因，也被证明在肠上皮发育中起作用。在转染细胞中的实验将确定这些转录因子的直接基因靶标。接下来，我将开发一种体内慢病毒siRNA系统，以确定Notch 1在肠道中的表型损失，并确定其在肠道分化层次中的位置。在目标3中，我将评估氧化偶氮甲烷在小鼠中产生肿瘤和异常隐窝病灶的作用，其中回肠和结肠含有不产生分泌细胞的Math 1-/-隐窝。从这些实验中产生的数据将形成一个独立的研究计划，以调查肠道发育和功能的基础。此外，这些目标代表了理解肠道生物学的不同方法，它们将相互通报，而不是相互依赖的成功。我将由肠道生物学和发育领域的领导者Susan Henning博士指导，并由相关领域的专家组成科学咨询委员会。PI拥有丰富的知识环境和广泛的资源。

DESCRIPTION (provided by applicant): In the intestinal epithelium, the balance between proliferation and apoptosis within the crypts of Lieberkuhn maintains villus height and crypt depth; disruption of this balance leads to cancer. Within the crypts, stem cells produce both absorptive and secretory cells. Several genes have been implicated in this process, but delineation of an ordered hierarchy for control of intestinal epithelial differentiation is lacking. One gene in this hierarchy, Math1, is required for secretory lineage development; mice lacking Math1, produce absorptive enterocytes but none of the secretory lineages. This proposal will test the hypotheses that Math1- dependent secretory lineages are required locally for proliferative homeostasis in the intestine, and that delineation of the genetic pathway for secretory lineage differentiation will clarify the mechanism of intestinal homeostasis. First I will characterize the genetic pathway for intestinal differentiation by microarray-based expression profiling of Math1 -/- crypts and crypts null for Gfi1, a putative Math1 target gene also shown to have a role in intestinal epithelial development. Experiments in transfected cells will define direct gene targets of these transcription factors. Next, I will develop an in vivo lentiviral-siRNA system to ascertain the phenotype of Notch1 loss in the intestine, and ascertain its position in the hierarchy of intestinal differentiation. In Aim 3, I will assess the effect of azoxymethane to produce tumors and aberrant crypt foci in mice in which the ileum and colon contains Math1 -/- crypts that produce no secretory cells. The data generated from these experiments will form the basis of an independent research program to investigate intestinal development and function. In addition, the aims represent distinct approaches to understanding intestinal biology, and they will inform one another while not being interdependent for success. I will be mentored by Dr. Susan Henning, a leader in the field of intestinal biology and development, with a scientific advisory committee composed of experts in relevant fields. A rich intellectual environment and extensive resources are available to the PI.