A novel serine protease inhibitor acts as an immunomodulatory switch while maintaining homeostasis.

Serine protease cascades boost immune responses while maintaining homeostasis. These crucial actions are intricately regulated by cognate serine protease inhibitors. However, the mechanism underlying such a dynamic immunomodulation during acute phase infection remains obscure, particularly where the pathogen's serine protease adds a new challenge to the host. Here, we found that infection of horseshoe crab, Carcinoscorpius rotundicauda, induced reciprocal profiles of CrSPI (serine protease inhibitor) and CrFurin (serine protease) with respect to their transcription and protein activities. Using recombinant rCrSPI, we explored its inhibitory activity against various microbial proteases and found it most efficacious against a model serine protease, subtilisin A. rCrSPI inhibited subtilisin at Ki 10(-9)M with a molar ratio of 1 rCrSPI:2 subtilisin. The rCrSPI also inhibited plasma CrFurin, suppressed subtilisin-mediated activation of prophenoloxidase (PPO) and interacted with complement C3. Taken together, CrSPI acts as a key immunomodulatory 'on-off' switch in a 2-way regulation of serine protease microbial subtilisin and host serine proteases (CrFurin and CrC3), thereby controlling immune responses involving the complements and the PPO-mediated antimicrobial activities, while maintaining homeostasis.