Cloning and function of sulfotransferase gene PmCHST1a in Pinctada martensii

Biomineralization is the process by which mineral crystals are deposited in an organized fashion in the matrix (either cellular or extracellular) of living organisms and the deposition of precise arrays of inorganic crystals in many organisms involves controlled nucleation at interfaces between the crystals and substrate macromolecules. Mollusc shells are used as a model for studying " organic-matrix-mediated” biomineralization, in which crystals are nucleated and grow in a pre-formed structural framework composed of proteins and polysaccharides. Acidic matrix macromolecules glycosaminoglycans were also associated with biological crystal growth and keratan sulfate widespread in animal tissues had a potential role in the process of biomineralization. Sulfotransferase plays a vital role in catalyzing the transfer of sulfonic acid groups in the processes of glycosaminoglycan synthesis, and glycosaminoglycans keratan sulfate with copious amounts of negatively charged sulfonic acid groups participate in the nucleation process of biomineralization. In this study, we cloned the sulfotransferase gene PmCHST1a (Pinctada martensiicarbohydrate sulfotransferase 1a) which participated in the synthesis of keratan sulfate and explored the function of sulfotransferase and keratan sulfate in the biomineralization. The full length sequence of PmCHST1a gene was obtained using the RACE technology, and we detected its expression pattern by real-time PCR. RNAi technique was used to study the potential functions of PmCHST1a in nacre formation, and we detected the effects to the synthesis of glycosaminoglycans keratan sulfate at the same time. The results showed that PmCHST1a, whose full length was 1 385 bp, encoded a protein of 366 amino acids. PmCHST1a carried with a typical Sulfotransferase-3 domain (Sulfotransfer-3 domain), a signal peptide and a transmembrane domain which made the protein located in the Golgi apparatus. The results of real-time PCR showed that PmCHST1a was highly expressed in the central zone of mantle. After the RNA interference experiments, the expression of PmCHST1a significantly decreased in the central zone of P. martensii, in union with the significant reduction of the concentration of glycosaminoglycans keratan sulfate in the extrapallial fluid. And the shell nacre crystallized irregularly compared with two control groups. These results showed that PmCHST1a may affect the formation of nacre through affecting the concentration of glycosaminoglycans keratan sulfate in the extrapallial fluid. This study provided the fundamental basis for further research of sulfotransferase and glycosaminoglycans in the formation of nacre.