Molecular regulation of gastric epithelial renewal

描述(由申请人提供)： 小鼠和人的胃上皮细胞经历了不断的更新。增殖异常不仅是胃病的共同特征，也是一般口服药物的共同特征。然而，人们对协调胃上皮细胞分裂、分化和迁移的分子机制知之甚少。目前的建议的目标是建立在先前工作的基础上，确定了胃上皮祖细胞(GEP)和胃酸分泌壁细胞(PC)的全球基因表达谱，PC是一种分化的胃谱系，被认为调节GEP的分化和分裂。GEP优先表达关键的躯体生长调节基因胰岛素样生长因子I(IGF-I)及其受体(IGF-ir)，而PC富含IGF隔离基因IGF结合蛋白2(IGFBP2)的表达。当前建议的第一个目标是检验PC使用IGFBP2来控制IGFS的生物利用度从而调节GEP增殖的假设。将产生其壁细胞将表现出转基因强制表达IGFBP2的小鼠。同时，通过产生携带IGFBP2等位基因的小鼠，并将其与实验室中已经产生的具有pc特异性cre重组酶表达的小鼠杂交，将有条件地去除IGFBP2。早期的工作还发现，GEP表达谱中20%的基因参与了mRNA的处理；许多果蝇的同源基因在胚胎发育过程中对极性的建立和细胞命运的决定至关重要。该提案的第二个目标将是确定RNA定位在胃上皮细胞更新中的作用。将开发工具来评估1)RNA定位蛋白(例如，GEP表达谱中的Mago-m和Y14基因)的亚细胞位置；2)与RNA定位装置相关的mRNA种类。为了识别候选的定位mRNAs，针对RNA定位蛋白的抗体将被用来免疫沉淀所有相关的mRNAs，并通过寡核苷酸阵列确定它们的身份。在果蝇中，其RNA定位的基因(例如，矮小和有毛的基因以及各种编码同源结构域的基因)对于分化发育中组织中的子细胞的命运至关重要。识别胃上皮细胞中的局部转录本可能会导致更好的分子图像，了解成年哺乳动物组织如何发生正常的更新，这反过来将有助于我们更好地表征胃上皮损伤反应，以及最终肿瘤(即异常)增殖是如何发展和可能被阻止的。

DESCRIPTION (provided by applicant): The gastric epithelium of mice and humans undergoes continuous renewal. Dysregulated proliferation is a common feature not only of gastric disease but of orally administered medication in general. Yet little is known about the molecular mechanisms that coordinate division, differentiation, and migration of gastric epithelial cells. The goal of the current proposal is to build on previous work that determined a global gene expression profile for both the gastric epithelial progenitor cell (GEP) and the acid-secreting parietal cell (PC), a differentiated gastric lineage that is thought to regulate GEP differentiation and division. GEPs preferentially express both the critical somatic growth regulating gene insulin-like growth factor I (IGF-I) and its receptor (IGF-Ir), whereas PCs are enriched for expression of the IGF-sequestering gene IGF binding protein 2 (IGFBP2). The first aim of the current proposal is to test the hypothesis that PCs use IGFBP2 to control bioavailability of IGFs thereby regulating GEP proliferation. Mice will be generated whose parietal cells will exhibit transgenic forced expression of IGFBP2. In parallel, IGFBP2 will be conditionally ablated by generating mice with floxed IGFBP2 alleles and crossing these to mice with PC-specific expression of cre recombinase that have already been generated in the lab. Earlier work also revealed that >20% of the genes in the GEP expression profile are involved in mRNA processing; many have Drosophila homologs critical for polarity establishment and cell fate determination during embryogenesis. The second aim of the proposal will be to characterize the role of RNA localization in gastric epithelial renewal. Tools will be developed to assess the subcellular location of 1) RNA localizing proteins (e.g., Mago-m and Y14, genes in the GEP expression profile); 2) mRNA species associated with the RNA localizing apparatus. To identify candidate localized mRNAs, antibodies against RNA localizing proteins will be used to immunoprecipitate all associated mRNAs and determine their identity by oligonucleotide array. In Drosophila, genes whose RNAs are localized (e.g., runt and hairy and various genes encoding homeodomains) are critical for differentiating cell fate of daughter cells in developing tissues. Identifying localized transcripts in the gastric epithelium may lead to a better molecular picture of how normal renewal occurs in adult mammalian tissue, which, in turn, would help us better characterize both the gastric epithelial injury response and, eventually, how neoplastic (i.e. abnormal) proliferation develops and might be prevented.