Cloning and functional characterization of PmΔ5FAD in pearl oyster Pinctada fucata martensii

Abstract

Pinctada fucata martensii produces high-quality pearls. However, excess immune and inflammatory response after grafting operation will lead to nucleus rejection, pearl sac formation failure, and death of the host pearl oyster. Polyunsaturated fatty acids (PUFAs) are critically important for its regulation function in inflammation and their synthesis process was limited by the related enzymes including fatty acid desaturase (FAD). In this study, FAD gene in the pearl oyster was characterized in P. f. martensii (PmΔ5FAD). We established that the full-length sequence of PmΔ5FAD contained a 1302-bp open reading frame that encoded a sequence of 433 amino acids. PmΔ5FAD domain analysis revealed the presence of a distinctive Cyt-b5 domain and a FA-desaturase domain that were highly similar to the Δ5FAD protein sequences identified in Haliotis discus hannai (69.25%) and Mimachlamys nobilis (62.96%). PmΔ5FAD expressed in all detected tissues. Functional analysis of PmΔ5FAD in recombinant Saccharomyces cerevisiae yeast showed the existence of Δ5-desaturation activity, which towards PUFA substrates and it efficiently desaturated exogenous PUFA C20:3n-6 to C20:4n-6 (ARA) with a desaturation conversion rate of 19.49%. Furthermore, PmΔ5FAD showed a significantly higher expression level on day 1 after the grafting operation, which may upregulate ARA synthesis in response to inflammatory and immune. These results provide important information concerning the function of FADs during pearl oyster PUFA biosynthesis and grafting operation.

Keywords

• PmΔ5FAD
• PUFA biosynthesis
• Pearl oyster
• Pinctada fucata martensii
• Grafting operation