Evolutionary perspective on regulation of Nramp1 expression in phagocytes

吞噬细胞中 Nramp1 表达调控的进化视角

• 批准号: 203297-2008
• 负责人: Cellier, Mathieu
• 金额: $ 1.68万
• 依托单位: Institut national de la recherche scientifique
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2008
• 资助国家: 加拿大
• 起止时间: 2008-01-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=392784
• 关键词: Evolutionary; perspective; regulation; Nramp1; expression

The role of the Natural resistance-associated macrophage protein (Nramp1) in host defense against microbial infections, including tuberculosis (TB) is widely accepted. This integral membrane protein limits access of phagocytosed microbes to intracellular nutrients, such iron and manganese. The activated form of Vitamin D3 (VD) is also known to enhance host innate resistance to TB, including treatments in solarium or using food supplements, which involvs increased production of antimicrobial peptides. Intracellular activation of Vitamin D3 precursor in human macrophages results in potent cell-autonomous anti Mycobacterium tuberculosis effect. We showed earlier that NRAMP1 expression is specifically induced by VD in macrophages, implying a possible role in VD antimicrobial effects. We began to delineate promoter regulatory sites and to identify bound factors. Achieving a comprehensive understanding of NRAMP1 regulation by VD will provide knowledge in terms of specific molecular mechanisms and possible avenues to stimulate host innate defenses. We thus propose to scan NRAMP1 gene and surrounding locus to identify all regulatory sites, positive or negative, that are mobilized during the development and maturation of professional phagocytes. We will seek optimal temporal frame for gene activity in time-course studies using inhibitors of chromosomal modeling. Gene activity-related DNA 'marks' and ('marked') bound factors, such as DNA packing histones or specific factors, will be purified using specific antibodies to precipitate fragments of fixed nuclear chromatin; bound DNA will be extracted and labeled before hybridization to a synthetic locus (high resolution tiling oligonucleotide micro-array). Conservation of the sites identified will be analyzed in the human genome to evaluate any significance of sequence polymorphisms, and genomes of other vertebrates to look for elements of a hypothetical ancestral regulatory system. This study will reveal the regulation in vivo of the endogenous NRAMP1 gene in human macrophages, establishing a knowledge basis for future innovative approaches aimed at stimulating host natural immunity and wound antimicrobial defense.

天然耐药相关巨噬细胞蛋白（Nramp1）在宿主防御微生物感染，包括结核病（TB）中的作用被广泛接受。这种完整的膜蛋白限制了被吞噬的微生物获得细胞内营养物质，如铁和锰。已知维生素D3 （VD）的活化形式可以增强宿主对结核病的先天抵抗力，包括日光疗法或使用食物补充剂，这涉及增加抗菌肽的产生。人巨噬细胞中维生素D3前体的胞内活化导致有效的细胞自主抗结核分枝杆菌作用。我们之前发现，巨噬细胞中的NRAMP1表达可被VD特异性诱导，这意味着它可能在VD抗菌作用中起作用。我们开始描绘启动子调控位点并确定结合因子。全面了解VD对NRAMP1的调控将为激发宿主先天防御提供具体的分子机制和可能的途径。因此，我们建议扫描NRAMP1基因及其周围位点，以确定在专业吞噬细胞发育和成熟过程中动员的所有阳性或阴性调控位点。我们将寻求基因活性的最佳时间框架在时间过程研究中使用抑制剂的染色体模型。基因活性相关的DNA“标记”和（“标记”）结合因子，如DNA包装组蛋白或特定因子，将使用特定抗体纯化，沉淀固定核染色质片段；结合的DNA将被提取并标记，然后杂交到合成位点（高分辨率平铺寡核苷酸微阵列）。这些位点的保存将在人类基因组中进行分析，以评估序列多态性的意义，并在其他脊椎动物的基因组中寻找假设的祖先调节系统的元素。本研究将揭示内源性NRAMP1基因在人巨噬细胞中的体内调控，为未来刺激宿主天然免疫和伤口抗菌防御的创新方法奠定知识基础。