Studies On Proteins And Genes Involved In The Shell Formation Of Pinctada Fucata

Biomineralization is the process that inorganic ions accumulated and deposited in the cellular or extracellular matrix. It is widely distributed in nature. Shell formation is one of the most typical biomineralization processes, in which the organic matrix, especially the matrix proteins, directs calcite or aragonite deposition at different positions and strictly controls the crystal nucleation, orientation and growth dynamics. Isolation of the proteins involving in shell formation and their genes would help to elucidate the molecular mechanism of shell formation, and also contribute to the development of new biomaterials and improvement of pearl production.A novel matrix protein, p10, was isolated from the nacreous layer of pearl oyster (Pinctada fucata) in this research. In vitro crystalline experiments showed that p10 could accelerate the nucleation of calcium carbonate crystals and induce the aragonite formation, suggesting a key role of p10 in polymorph control. Cellular assays showed that p10 could stimulate the differentiation of fibroblasts MRC-5 and pre-osteoblasts MC3T3-E1, suggesting that p10 might also influence the mineralgenic cells. Taken together, p10 seems to be a multi-functional matrix protein, which plays important roles in nacre biomineralization.Besides isolating the matrix proteins from the shell directly, three kinds of genes encoding tyrosinase OT47, matrix protein KRMP family and myosin essential light chain respectively, which were all involved in shell formation, were also cloned from the mantle of P. fucata, and their tissue distribution were investigated. Tyrosinase OT47 was expressed specifically in the outer epithelial cells of the middle mantle fold, suggesting that it contributes to periostracum formation and the periostracum secretion model of P. fucata is different from previous opinion. The KRMP family contained a putative signal peptide, which shared similarity with other known prismatic layer matrix proteins, and was expressed specifically in the epithelial cells of the outer mantle fold, suggesting KRMP is a prismatic layer matrix protein family. It was also hypothesized via sequence analysis that KRMP may anchor acidic proteins, which was thought to direct the CaCO3 crystals nucleation, to the matrixframework. The sequence and expression analysis of myosin essential light chain gene may also help to further understand the Ca2+ metabolism and the secretion of shell matrix proteins in P. fucata.