Molecular cloning, expression and functional analysis of B-cell activating factor (BAFF) in yellow grouper, Epinephelus awoara

• We first report the cloning and characterization of the yellow grouper BAFF.
• EasBAFF was efficiently expressed and purified.
• WST-8 assay indicated that EasBAFF could promote the survival/proliferation of yellow grouper splenic lymphocytes.
• The role of BAFF may be useful for understanding the anti-bacteria immunity in fish.

B cell activating factor (BAFF), a ligand belonging to the tumor necrosis factor (TNF) family is critical to B cell survival, proliferation, maturation and immunoglobulin secretion. In this study, the yellow grouper (Epinephelus awoara) BAFF (designated EaBAFF) gene was cloned using RT-PCR and RACE (rapid amplification of cDNA ends) techniques. The full-length EaBAFF was 1442 bp and contained an open reading frame of 780 bp encoding a putative protein of 259 amino acids. Amino acids sequence comparison indicated that EaBAFF possessed the TNF signature. The soluble BAFF (EasBAFF) had been cloned into pET28a. SDS-PAGE and Western blotting analysis confirmed that the soluble fusion protein His-EasBAFF was efficiently expressed in Escherichia coli BL21 (DE3). In vitro, the WST-8 assay indicated that EasBAFF was not only able to promote the survival/proliferation of yellow grouper splenic lymphocytes but also able to promote the survival/proliferation of mouse splenic B cells. Our findings may provide valuable information for research into the immune system of E. awoara and EasBAFF may serve as a potential immunologic factor for enhancing immunological efficacy in fish.

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