CDK5RAP3, a key defender of udder, modulates NLRP3 inflammasome activation by regulating autophagolysosome degradation in S. agalactiae-infected mastitis

Streptococcus agalactiae, as one of the main pathogens of clinical and subclinical mastitis, affects animal welfare and leads to huge economic losses to farms due to the sharp decline in milk yield. However, both the real pathogenic mechanisms of S. agalactiae-induced mastitis and the regulator which controls the inflammation and autophagy are largely unknown. Served as a substrate of ubiquitin-like proteins of E3 ligase, CDK5RAP3 is widely involved in the regulation of multiple signaling pathways. Our findings revealed that CDK5RAP3 was significantly down-regulated in mastitis infected by S. agalactiae. Surprisingly, inflammasome activation was triggered by CDK5RAP3 knockdown: up-regulated NLRP3, IL1β and IL6, and cleaved caspase1 promoting by NF-κB, thereby resulting in pyroptosis. Additionally, the accumulation of autophagy markers (LC3B and p62) after CDK5RAP3 knockdown suggested that the autophagolysosome degradation pathway was inhibited, thereby activating the NF-κB pathway and NLRP3 inflammasome. Hence, our findings suggest that downregulation or ablation of CDK5RAP3 inhibits autophagolysosome degradation, causes inflammation by activating the NF-κB /NLRP3 inflammasome, and triggers cell death. In conclusion, CDK5RAP3 holds the key to understanding the interaction between autophagy and immune responses, its anti-inflammatory role in this study will throw new light on the clinical drug discovery to cure S. agalactiae mastitis.