Characterization of the mechanism regulating P2X7 expression and functions in intestinal epithelial cells.

肠上皮细胞中 P2X7 表达和功能调节机制的表征。

• 批准号: 327128-2013
• 负责人: Gendron, FernandPierre
• 金额: $ 2.62万
• 依托单位: Université de Sherbrooke
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2014
• 资助国家: 加拿大
• 起止时间: 2014-01-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=548986
• 关键词: Characterization; mechanism; regulating; P2X7; expression

The formation of a mature and functional intestinal epithelium requires the integration of an array of extracellular messages by immature epithelial cells to give rise to differentiated and functional intestinal epithelial cells (IECs). These messages lead to expression of genes required for IECs differentiation by modifying DNA regulatory sequences and by the recruitment of specific proteins, namely histones and transcription factors, to allow gene expression. We showed that adenosine triphosphate transmits such messages by activating P2X7, a cell surface receptor expressed by differentiated IECs. Indeed, the absence of P2X7 expression in mice (P2X7-/-) results in aberrant epithelium structure. In this research program, we will determine how P2X7 expression is regulated in IECs and how this receptor contributes to the establishment of a functional intestine. To unravel the regulatory mechanisms regulating P2X7 expression and functions, we will use Caco-2, a human epithelial cell model recapitulating IEC differentiation, IECs isolated from P2X7-/- mice and normal human intestinal biopsies. Modification to the DNA regulatory sequences controlling P2X7 expression will be characterized by analyzing the DNA sequence and by identifying histones and transcription factors associated to this region. To characterize P2X7 roles in intestinal epithelium structure, we will use classical approaches, such as the characterization of cell-to-cell junction, cell proliferation rate and cell determination analysis, in P2X7-/- mice and, in a second time, we will use an unbiased approach to define the global impact of P2X7 expression in IECs function by profiling the entire genome (transcriptome) using high-throughput RNA sequencing. Finally, we will validate these findings in the Caco-2 human model of intestinal epithelial cell. The expected results from this research program will define, for the first time, the molecular and cellular mechanisms governing the regulation of P2X7 expression and its putative role in IECs. Given the polymorphisms found between humans resulting in loss- or gain-of-receptor functions, our results will bring a new understanding on the role of this receptor in human physiology.

成熟和功能性肠上皮的形成需要由未成熟的上皮细胞整合一系列细胞外信息，以引起分化和功能性的肠上皮细胞（IEC）。这些信息通过修饰DNA调节序列以及募集特定蛋白质（即组蛋白和转录因子）来允许基因表达，从而导致IEC分化所需的基因。我们表明，三磷酸腺苷通过激活P2X7（由分化IEC表达的细胞表面受体）激活P2X7来传播此类信息。实际上，小鼠（P2X7 - / - ）中缺乏P2X7表达会导致异常上皮结构。在该研究计划中，我们将确定如何在IEC中调节P2X7的表达，以及该受体如何促进功能性肠的建立。为了阐明调节P2X7表达和功能的调节机制，我们将使用CACO-2，一种概括IEC分化的人类上皮细胞模型，从P2X7 - / - 小鼠和正常的人类肠生活检中分离出的IEC。对控制P2X7表达的DNA调节序列的修改将以分析DNA序列和鉴定与该区域相关的组蛋白和转录因子来表征。 To characterize P2X7 roles in intestinal epithelium structure, we will use classical approaches, such as the characterization of cell-to-cell junction, cell proliferation rate and cell determination analysis, in P2X7-/- mice and, in a second time, we will use an unbiased approach to define the global impact of P2X7 expression in IECs function by profiling the entire genome (transcriptome) using high-throughput RNA sequencing.最后，我们将在肠上皮细胞的CACO-2人类模型中验证这些发现。该研究计划的预期结果将首次定义有关P2X7表达调节及其在IEC中的作用的分子和细胞机制。鉴于人类之间发现的多态性导致受损失或受体功能的损失，我们的结果将对该受体在人类生理学中的作用有了新的了解。