Mechanism of Activation of Innate Immunity by ISS-DNA

ISS-DNA 激活先天免疫的机制

• 批准号: 6885363
• 负责人: Wen-Ming Chu
• 金额: $ 27.48万
• 依托单位: BROWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-15 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6885363
• 关键词: bacterial DNA; biological signal transduction; immunoregulation; laboratory mouse; leukocyte activation /transformation; macrophage; microorganism immunology; protein kinase; toll like receptor

DESCRIPTION (provided by applicant): Innate immunity enables an organism to respond rapidly to invading microorganisms. To do this, innate immune system uses a variety of receptors or intracellular proteins that can recognize a microbial pathogen by the motifs pathogen-associated molecular patterns (PAMPs). Pathogen-associated motifs include mannans in the yeast cell wall, formylated peptides and various bacterial cell-wall components such as lipopolysaccharide (LPS), lipopeptides, peptidoglycans and teichoic acids. Bacterial DNA is also a potent inducer of innate immunity. Bacterial DNA contains increased frequency of CpG dinucleotide motifs in particular sequence contexts (PuPuCpGPyPy), which are greatly reduced in frequency in the mammalian genome, and where it occurs, the cytosine commonly is methylated. DNA containing unmethylated CpG motifs is immunostimulatory and able to activate cellular component of innate immunity, including macrophages and dendritic cells (DC) as well as B cells. Thus, DNA containing unmethylated CpG motif in sequences of PuPuCpGPyPy is also called ISS-DNA. Activation of immune cells by ISS-DNA appears to require uptake into the cells by receptor- or pinocytosis-mediated endocytosis and endosome acidification, suggesting that the recognition structure is either endosome-associated or cytoplasmic. Recent studies suggested that Toll-like receptor (TLR) 9 and adapter protein MyD88 are required for ISS-DNA to induce immune cells to produce interlukin-12, and for activation of NF-kB by ISS-ODN. The studies from our group suggested that catalytic subunit of DNA-dependent protein kinase (DNAPKcs) and IkB kinase (IKK) are required for the activation of innate immunity by ISS-DNA. ISS-DNA activates DNA-PKcs, which in turn activates IKK, leading to NF-kB activation. Null mutations in either the DNA-PKcs- or IKKb gene were found to selectively impair the ability of mice or their macrophages, to respond to ISS-DNA with inducible cytokine gene expression. TLR9 is on cell membrane while DNA-PKcs is in cytoplasm and nucleus. The connection between TLR9 and DNA-PKcs is not apparent. To further understand the molecular mechanisms of activation of innate immunity by ISS-DNA, we will investigate the relationship between TLR9-signaling pathway and DNA-PK by using biochemical and genetic means. Moreover, we will investigate if the regulatory subunits of DNA-PK, Ku70 and Ku80, and their associated factors are required for the innate response to ISS-DNA. Furthermore, we will identify new molecules that are required for activation of innate immunity by ISS-DNA.

描述（由申请人提供）：先天免疫使生物体能够 对入侵的微生物迅速作出反应。为了做到这一点，先天免疫系统 使用多种受体或细胞内蛋白质，可以识别 微生物病原体的基序病原体相关分子模式 （PAMP）。病原体相关基序包括酵母细胞壁中的甘露聚糖， 甲酰化肽和各种细菌细胞壁组分， 脂多糖（LPS）、脂肽、肽聚糖和磷壁酸。 细菌DNA也是先天免疫的有效诱导剂。 细菌DNA中含有增加频率的CpG二核苷酸基序， 特定序列上下文（PuPuCpGPyPy），这在 在哺乳动物基因组中的频率，以及它发生的地方，胞嘧啶通常 是甲基化的。含有未甲基化CpG基序的DNA是免疫刺激性的， 能够激活先天免疫的细胞成分，包括巨噬细胞 和树突细胞（DC）以及B细胞。因此，含有未甲基化的DNA PuPuCpGPyPy序列中的CpG基序也称为ISS-DNA。激活 免疫细胞的ISS-DNA似乎需要吸收到细胞的受体- 或胞饮介导的内吞作用和内体酸化，这表明， 识别结构是内体相关的或细胞质的。最近 研究表明，Toll样受体（TLR）9和衔接蛋白MyD 88是 ISS-DNA诱导免疫细胞产生白细胞介素-12所需， 通过ISS-ODN激活NF-kB。我们小组的研究表明， DNA依赖性蛋白激酶催化亚基（DNAPKcs）和IkB激酶 (IKK)是通过ISS-DNA激活先天免疫所必需的。ISS-DNA 激活DNA-PKcs，继而激活IKK，导致NF-κ B激活。 在DNA-PKcs-或IKKb基因的突变被发现选择性地 损害小鼠或其巨噬细胞对ISS-DNA的反应能力， 可诱导细胞因子基因表达。 TLR 9定位于细胞膜上，DNA-PKcs定位于细胞质和细胞核内。的 TLR 9和DNA-PKcs之间的联系并不明显。进一步了解 ISS-DNA激活先天免疫的分子机制，我们将 探讨TLR 9信号通路与DNA-PK的关系， 生物化学和遗传学手段。此外，我们将调查监管机构是否 需要DNA-PK、Ku 70和Ku 80亚基及其相关因子 对ISS-DNA的先天反应。此外，我们将识别新的分子 是通过ISS-DNA激活先天免疫所必需的。