The dual role of eppin in immunity and biomineralization during nacreous layer formation in mollusks

Abstract

The presence of protease inhibitors in a carbonated shell is intriguing and opens questions about their potential role in shell biomineralization. Our previous proteome study detected an epididymal protease inhibitor (eppin) which was strongly expressed in pearl nacre. Herein, we obtained the full-length complementary DNA sequence of eppin from Hyriopsis cumingii. The expression of eppin was localized to the dorsal epithelial cells of the mantle pallium and mantle center and significantly increased during pearl nacre deposition and shell nacreous layer regeneration. These results indicated that eppin is essential for nacre biomineralization. The expression of eppin was significantly increased at 3 h after Staphylococcus aureus challenge. GST-eppin could significantly inhibit the growth of S. aureus and effectively inhibit chymotrypsin activity. Inhibition of eppin in extrapallial fluid induced haphazard distribution of nanoscale grains and the destruction of the organic framework of the nacreous layer. These results indicated that eppin had a long-lasting antibacterial activity and provided a matrix protecting system that prevented extracellular degradation of matrix proteins. In addition, GST-eppin affected morphology regulation and induced the acceleration of calcium carbonate deposition. These results added to our understanding of the mode of action of protease inhibitors and their roles in shell biomineralization.