Nod1: An Apaf-like Activator of Apoptosis and NF-kB

Nod1：一种类似 Apaf 的细胞凋亡和 NF-kB 激活剂

• 批准号: 6757253
• 负责人: NAOHIRO INOHARA
• 金额: $ 20万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6757253
• 关键词: antigen antibody reaction; apoptosis; bacteria infection mechanism; bacterial disease; bacterial genetics; bacterial polysaccharides; cell membrane; immunoprecipitation; immunoregulation; laboratory mouse; lipopolysaccharides; monocyte; neutrophil; nuclear factor kappa beta; nuclear receptors; posttranslational modifications; protein kinase; protein protein interaction; protein structure function; thin layer chromatography

The immune response to microbial pathogens is initiated by recognition of specific pathogen components by host cells both at the cell surface and in the cytosol. While the response triggered by pathogen products at the surface of immune cells is well characterized, that initiated in the cytosol is poorly understood. Nod1 is a member of a growing family of proteins with structural homology to apoptosis regulators Apaf-1/Ced-4 and plant disease resistant R gene products. Nod1 promotes apoptosis when overexpressed in cells, but unlike Apaf-1, it induces NF-kappaB activation. NF-KappaB activation induced by Nod1 is mediated by the association of the CARD of Nod1 with the corresponding CARD of RICK, a protein kinase that activates NF-kappaB. Analyses with wild-type (wt) and mutant forms of both Nod1 and RICK have suggested that Nod1 and RICK act in the same pathway of NF-kappaB activation, where RICK functions as a downstream mediator of Nod1 signaling. Nod1 self-associates through its nucleotide-binding domain (NBD) and Nod1 oligomerization promotes proximity of RICK molecules and NF-kappaB activation. Like Nod1, intracellular plant R proteins contain N-terminal effector domains linked to a NBD and multiple leucine-rich repeats (LRRs) located C-terminally of the NBD. The LRRs of R proteins are highly diverse and appear to be involved in the recognition of a wide array of pathogen components. Remarkably, we find that bacterial lipopolysacharides (LPS), but not other pathogen components, induced TLR4- and MyD88- independent NF-kappaB activation in human embryonic kidney 293T cells expressing trace amounts of Nod1. Like plant disease resistant R proteins, the LRRs of Nod1 were required for LPS-induced NF-kappaB activation. Furthermore, a LPS binding activity could be co-immunopurified with Nod1 from cytosolic extracts. Our hypothesis is that Nod1 is a mammalian counterparts of plant R gene products that may function as a cytosolic receptor for pathogen components derived from invading bacteria. We propose three Specific Aims to understand the biochemical mechanism by which bacterial LPS and Nod1 interact and to determine the role of Nod1 in the response to bacterial LPS in vivo.

针对微生物病原体的免疫反应是通过宿主细胞在细胞表面和细胞质中识别特定病原体成分​​而启动的。虽然免疫细胞表面病原体产物引发的反应已得到很好的表征，但对细胞质中引发的反应却知之甚少。 Nod1 是一个不断增长的蛋白质家族的成员，与凋亡调节因子 Apaf-1/Ced-4 和植物抗病 R 基因产物具有结构同源性。 Nod1 在细胞中过度表达时会促进细胞凋亡，但与 Apaf-1 不同的是，它会诱导 NF-kappaB 激活。 Nod1 诱导的 NF-KappaB 激活是通过 Nod1 的 CARD 与 RICK（一种激活 NF-kappaB 的蛋白激酶）相应 CARD 的结合介导的。 对野生型 (wt) 和突变型 Nod1 和 RICK 的分析表明，Nod1 和 RICK 在 NF-kappaB 激活的同一途径中发挥作用，其中 RICK 充当 Nod1 信号传导的下游介质。 Nod1 通过其核苷酸结合域 (NBD) 自关联，Nod1 寡聚化促进 RICK 分子的接近和 NF-kappaB 激活。 与 Nod1 一样，细胞内植物 R 蛋白包含与 NBD 连接的 N 端效应结构域和位于 NBD C 端的多个富含亮氨酸重复序列 (LRR)。 R 蛋白的 LRR 高度多样化，并且似乎参与多种病原体成分​​的识别。值得注意的是，我们发现，在表达微量Nod1的人胚胎肾293T细胞中，细菌脂多糖（LPS）而非其他病原体成分​​诱导TLR4和MyD88独立的NF-κB激活。 与植物抗病 R 蛋白一样，Nod1 的 LRR 是 LPS 诱导 NF-κB 激活所必需的。 此外，LPS 结合活性可以与胞浆提取物中的 Nod1 共同免疫纯化。 我们的假设是，Nod1 是植物 R 基因产物的哺乳动物对应物，可能充当来自入侵细菌的病原体成分​​的胞质受体。 我们提出了三个具体目标，以了解细菌 LPS 和 Nod1 相互作用的生化机制，并确定 Nod1 在体内细菌 LPS 反应中的作用。