1. Molecular structure, expression, and bioactivity of B-cell-activating factor of the TNF family (BAFF) and its receptor BAFF-R in cats (Felis catus)
- Author
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Xiaolong Wu, Zhiheng Wei, Jianfeng Li, Lei Ma, Zhiguo Wang, Jia-Xin Zhang, Hongzhen Liu, Ming Sang, and Zhang Shuangquan
- Subjects
0301 basic medicine ,Staphylococcus aureus ,DNA, Complementary ,Cell Survival ,Immunology ,Cross Reactions ,Green fluorescent protein ,03 medical and health sciences ,Mice ,Open Reading Frames ,0302 clinical medicine ,Complementary DNA ,B-Cell Activating Factor ,Escherichia coli ,Animals ,Amino Acid Sequence ,Cloning, Molecular ,B-cell activating factor ,Receptor ,Molecular Biology ,Furin ,Mammals ,B-Lymphocytes ,biology ,Base Sequence ,Molecular Structure ,Chemistry ,Fusion protein ,Molecular biology ,Recombinant Proteins ,Reverse transcription polymerase chain reaction ,Transmembrane domain ,030104 developmental biology ,Immunoglobulin M ,biology.protein ,Cats ,Spleen ,030215 immunology ,B-Cell Activation Factor Receptor - Abstract
B-cell survival depends on signals induced by binding of B-cell activating factor (BAFF) to its receptor (BAFF-R). In this study, the full-length cDNAs of cat BAFF (cBAFF) and BAFF-R (cBAFF-R) were amplified from the spleen by reverse transcription PCR. The open reading frame of cBAFF cDNA encodes a protein of 285 amino acids containing a predicted transmembrane domain and a furin protease cleavage site, similar to mammalian, avian, and reptile BAFFs. The cBAFF-R gene encodes a 189 amino acid protein. Real-time quantitative PCR analyses revealed that the two genes are predominantly expressed in the spleen. csBAFF, EGFP/csBAFF, and cBAFF-R were efficiently expressed in Escherichia coli BL21 (DE3), as confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting analyses. After purification, the EGFP/csBAFF fusion protein showed a fluorescence spectrum similar to that of EGFP. Confocal laser scanning microscopy showed that EGFP/csBAFF bound to its receptor. In vitro, csBAFF promoted the survival of cat and mouse splenic B cells with/without a priming agent (Staphylococcus aureus Cowan 1, SAC) or anti-mouse IgM. Furthermore, it stimulated the survival of mouse B cells, similar to msBAFF. Recombinant cBAFF-R blocked the function of sBAFF in vitro. These findings indicate that csBAFF plays an important role in the survival of cat B cells and has functional cross reactivity between cats and other mammals, and suggest a role for the BAFF-BAFF-R system in regulating B-cell survival. Therefore, BAFF and BAFF-R show promise for enhancing the immune systems of animals.
- Published
- 2019