Molecular cloning, expression and functional analysis of B-cell activating factor (BAFF) in yellow grouper, Epinephelus awoara.
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 1442bp and contained an open reading frame of 780bp 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.
Admission and Employment Office, JiangSu Second Normal University, Nanjing 210013, China.
Wen
W
Xiao
Nanjing Maternal and Child Health Medical Institute, Nanjing Medical University Affiliated Nanjing Maternal and Child Health Hospital, Nanjing, China.
Wei
W
Long
Admission and Employment Office, JiangSu Second Normal University, Nanjing 210013, China.
Gui-you
GY
Liu
Admission and Employment Office, JiangSu Second Normal University, Nanjing 210013, China.
Chun-liang
CL
Sui
Nanjing Maternal and Child Health Medical Institute, Nanjing Medical University Affiliated Nanjing Maternal and Child Health Hospital, Nanjing, China.
Xi-rong
XR
Guo
Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing, China.
Aiying
A
Tian
Nanjing Maternal and Child Health Medical Institute, Nanjing Medical University Affiliated Nanjing Maternal and Child Health Hospital, Nanjing, China.
Chen-bo
CB
Ji
Nanjing Maternal and Child Health Medical Institute, Nanjing Medical University Affiliated Nanjing Maternal and Child Health Hospital, Nanjing, China. Electronic address: cuixianwei198615@163.com.
Xian-wei
XW
Cui
Jiangsu Province Key Laboratory for Molecular and Medical Biotechnology, Life Sciences College, Nanjing Normal University, Nanjing, China. Electronic address: shuangquan_z@yahoo.com.
Shuang-Quan
SQ
Zhang
D000595:Amino Acid Sequence; D000818:Animals; D053264:B-Cell Activating Factor / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D001402:B-Lymphocytes / Q000378:metabolism; D015153:Blotting, Western; D003001:Cloning, Molecular; D029941:Fish Proteins / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010473:Perciformes / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D011485:Protein Binding; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000166:cytology; D059467:Transcriptome
D053264:B-Cell Activating Factor; D029941:Fish Proteins
B-cell survival; BAFF; RACE; Yellow grouper (E. awoara)