NLRP3 inflammasome activation is pivotal for cytokine secretion and pyroptosis in response to diverse stimuli, playing a crucial role in innate immunity. While extensively studied in mammals, the regulatory mechanisms governing NLRP3 activation in non-mammalian vertebrates remain largely unexplored. Teleosts, as basal vertebrates, represent an ideal model for exploring the evolutionary trajectory of inflammasome regulation. In this study, ABE assays, confocal microscopy, and biochemical analyses were applied to systematically characterize the mechanisms underlying NLRP3 inflammasome in teleosts, using large yellow croakers ( Larimichthys crocea, Lc) and zebrafish ( Danio rerio, Dr) as representative models. Our findings revealed a previously unrecognized palmitoylation-dependent regulatory mechanism essential for teleost NLRP3 activation. Specifically, zDHHC18-mediated palmitoylation at a teleost-specific cysteine residue (C946 in LcNLRP3, C1037 in DrNLRP3) was required for the translocation of NLRP3 to the dispersed trans-Golgi network, facilitating its subsequent recruitment to the microtubule-organizing center. This membrane trafficking was crucial for inflammasome assembly and downstream inflammatory responses. These findings provide new insights into the distinct regulatory mechanisms of NLRP3 activation in teleosts, highlighting an evolutionary divergence that contributes to innate immunity adaptation in early vertebrates.
NLRP3炎症小体的激活在应对病原感染等应激条件下的细胞因子分泌和焦亡过程中起着至关重要的作用，是固有免疫反应中的关键环节。尽管在哺乳动物中已有广泛研究，但非哺乳类脊椎动物中NLRP3炎性小体激活的调控机制仍然较少探索。作为脊椎动物早期分支物种，硬骨鱼为研究炎症小体调节的进化提供了独特的视角。该研究采用棕榈酸生物素交换（ABE）法、共聚焦显微镜和生化分析等方法，以大黄鱼（ Larimichthys crocea， Lc）和斑马鱼（ Danio rerio， Dr）为代表性模型，系统地表征了硬骨鱼NLRP3炎症小体活化的调节机制。研究发现了硬骨鱼NLRP3特异位点的棕榈酰化修饰介导的炎症小体激活机制。具体来说，我们发现zDHHC18介导了硬骨鱼NLRP3特有的半胱氨酸位点（ LcNLRP3中的C946、 DrNLRP3中的C1037）的棕榈酰化，这一过程对NLRP3向解聚的反式高尔基囊膜（dTGN）的转运以及随后在微管组织中心（MTOC）的定位至关重要，并对后续炎症小体的组装和炎症反应的活化具有重要作用。该研究为硬骨鱼NLRP3激活的独特调控机制提供了新的见解，揭示了免疫调节机制在进化过程中的差异，并加深了我们对固有免疫适应的理解。.
Keywords: Inflammasome activation; NLRP3; Palmitoylation; Teleost; zDHHC18.