Inflammasomes are signalling platforms that are assembled in response to infection or sterile inflammation by cytosolic pattern recognition receptors. The consequent inflammasome-triggered caspase-1 activation is critical for the host defence against pathogens. During infection, NLRP3, which is a pattern recognition receptor that is also known as cryopyrin, triggers the assembly of the inflammasome-activating caspase-1 through the recruitment of ASC and Nek7. The activation of the NLRP3 inflammasome is tightly controlled both transcriptionally and post-translationally. Despite the importance of the NLRP3 inflammasome regulation in autoinflammatory and infectious diseases, little is known about the mechanism controlling the activation of NLRP3 and the upstream signalling that regulates the NLRP3 inflammasome assembly. We have previously shown that the Rho-GTPase-activating toxin from Escherichia coli cytotoxic necrotizing factor-1 (CNF1) activates caspase-1, but the upstream mechanism is unclear. Here, we provide evidence of the role of the NLRP3 inflammasome in sensing the activity of bacterial toxins and virulence factors that activate host Rho GTPases. We demonstrate that this activation relies on the monitoring of the toxin’s activity on the Rho GTPase Rac2. We also show that the NLRP3 inflammasome is activated by a signalling cascade that involves the p21-activated kinases 1 and 2 (Pak1/2) and the Pak1-mediated phosphorylation of Thr 659 of NLRP3, which is necessary for the NLRP3–Nek7 interaction, inflammasome activation and IL-1β cytokine maturation. Furthermore, inhibition of the Pak–NLRP3 axis decreases the bacterial clearance of CNF1-expressing UTI89 E. coli during bacteraemia in mice. Taken together, our results establish that Pak1 and Pak2 are critical regulators of the NLRP3 inflammasome and reveal the role of the Pak–NLRP3 signalling axis in vivo during bacteraemia in mice.

炎症小体是信号平台，通过胞质模式识别受体响应感染或无菌炎症而组装。随后的炎症小体触发的 caspase-1 激活对于宿主防御病原体至关重要。在感染过程中，NLRP3（一种模式识别受体，也称为冷吡蛋白）通过招募 ASC 和 Nek7 来触发炎症小体激活 caspase-1 的组装。 NLRP3 炎症小体的激活在转录和翻译后均受到严格控制。尽管 NLRP3 炎症小体调节在自身炎症和感染性疾病中很重要，但人们对控制 NLRP3 激活的机制以及调节 NLRP3 炎症小体组装的上游信号传导知之甚少。我们之前已经证明来自大肠杆菌细胞毒性坏死因子-1（CNF1）的Rho-GTP酶激活毒素可以激活caspase-1，但上游机制尚不清楚。在这里，我们提供了 NLRP3 炎症小体在感知细菌毒素和激活宿主 Rho GTP 酶的毒力因子活性中的作用的证据。我们证明这种激活依赖于对 Rho GTPase Rac2 上毒素活性的监测。我们还表明，NLRP3 炎症小体是由信号级联激活的，该信号级联涉及 p21 激活的激酶 1 和 2 (Pak1/2) 以及 Pak1 介导的 NLRP3 Thr 659 的磷酸化，这对于 NLRP3-Nek7 相互作用是必需的，炎症小体激活和 IL-1β 细胞因子成熟。此外，抑制 Pak-NLRP3 轴会降低小鼠菌血症期间表达 CNF1 的 UTI89大肠杆菌的细菌清除率。 综上所述，我们的结果证实 Pak1 和 Pak2 是 NLRP3 炎症小体的关键调节因子，并揭示了 Pak-NLRP3 信号轴在小鼠菌血症期间体内的作用。