Department of Pathology, Proteo-Science Center and Graduate School of Medicine, Ehime University, Toon, Ehime, Japan.
Department of Hepatobiliary Pancreatic and Transplantation Surgery, Graduate School of Medicine, Ehime University, Toon, Ehime, Japan.
J Immunol. 2024 Jul 1;213(1):86-95. doi: 10.4049/jimmunol.2300175.
The nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3, also called cryopyrin) inflammasome is an intracellular innate immune complex, which consists of the pattern-recognition receptor NLRP3, the adaptor apoptosis-assciated speck-like protein containing a caspase recruitment domain, and procaspase-1. Aberrant activation of the NLRP3 inflammasome causes an autoinflammatory disease called cryopyrin-associated periodic syndrome (CAPS). CAPS is caused by gain-of-function mutations in the NLRP3-encoding gene CIAS1; however, the mechanism of CAPS pathogenesis has not been fully understood. Thus, unknown regulators of the NLRP3 inflammasome, which are associated with CAPS development, are being investigated. To identify novel components of the NLRP3 inflammasome, we performed a high-throughput screen using a human protein array, with NLRP3 as the bait. We identified a NLRP3-binding protein, which we called the cryopyrin-associated nano enhancer (CANE). We demonstrated that CANE increased IL-1β secretion after NLRP3 inflammasome reconstitution in human embryonic kidney 293T cells and formed a "speck" in the cytosol, a hallmark of NLRP3 inflammasome activity. Reduced expression of endogenous CANE decreased IL-1β secretion upon stimulation with the NLRP3 agonist nigericin. To investigate the role of CANE in vivo, we developed CANE-transgenic mice. The PBMCs and bone marrow-derived macrophages of CANE-transgenic mice exhibited increased IL-1β secretion. Moreover, increased autoinflammatory neutrophil infiltration was observed in the s.c. tissue of CANE-transgenic versus wild-type mice; these phenotypes were consistent with those of CAPS model mice. These findings suggest that CANE, a component of the NLRP3 inflammasome, is a potential modulator of the inflammasome and a contributor to CAPS pathogenesis.
核苷酸结合寡聚化结构域样受体家族富含pyrin 结构域蛋白 3（NLRP3，也称为冷吡啉）炎症小体是一种细胞内先天免疫复合物，由模式识别受体 NLRP3、衔接凋亡相关斑点样蛋白含有半胱氨酸蛋白酶募集结构域和前胱天蛋白酶-1 组成。NLRP3 炎症小体的异常激活导致一种称为冷吡啉相关周期性综合征（CAPS）的自身炎症性疾病。CAPS 是由 NLRP3 编码基因 CIAS1 的功能获得性突变引起的；然而，CAPS 发病机制的机制尚未完全理解。因此，正在研究与 CAPS 发展相关的 NLRP3 炎症小体的未知调节剂。为了鉴定 NLRP3 炎症小体的新成分，我们使用人蛋白阵列作为诱饵进行了高通量筛选。我们鉴定出一种与 NLRP3 结合的蛋白，我们称之为冷吡啉相关纳米增强子（CANE）。我们证明，在人胚肾 293T 细胞中重建 NLRP3 炎症小体后，CANE 可增加 IL-1β 的分泌，并在细胞质中形成“斑点”，这是 NLRP3 炎症小体活性的标志。内源性 CANE 表达减少会降低 NLRP3 激动剂 Nigericin 刺激后的 IL-1β 分泌。为了研究 CANE 在体内的作用，我们开发了 CANE 转基因小鼠。CANE 转基因小鼠的 PBMC 和骨髓来源的巨噬细胞表现出增加的 IL-1β 分泌。此外，与野生型小鼠相比，CANE 转基因小鼠的皮肤下组织中观察到嗜中性粒细胞浸润增加的自发性炎症；这些表型与 CAPS 模型小鼠的表型一致。这些发现表明，NLRP3 炎症小体的组成部分 CANE 是炎症小体的潜在调节剂，也是 CAPS 发病机制的贡献者。
