Cloning, characterization, and expression analysis of orange-spotted grouper (Epinephelus coioides) ILF2 gene (EcILF2).
Interleukin-2 enhancer binding factor 2 (ILF2), also named as nuclear factor 45 (NF45), plays important roles in regulating interleukin-2 expression in mammals. In the present study, a novel ILF2 gene (designated EcILF2) was cloned and well characterized from orange-spotted grouper, Epinephelus coioides. The full-length EcILF2 cDNA is composed of 1544 bp and encodes a polypeptide of 387 amino acids with 98% identity to ILF2 of Atlantic salmon. The genomic DNA of EcILF2 consists of 14 exons and 13 introns, with a length of approximately 6.9 kb. EcILF2 contains two conserved domains including an RGG-rich single-stranded RNA-binding domain and a DZF zinc-finger nucleic acid binding domain. Recombinant EcILF2 was expressed in Escherichia coli BL21 (DE3) and purified for mouse anti-EcILF2 serum preparation. Subcellular localization analysis revealed that EcILF2 was distributed predominantly in the nucleus. Realtime quantitative PCR (RT-qPCR) analysis revealed a broad expression of EcILF2, with a relative strong expression in skin, liver, brain, head kidney and spleen. The expression of EcILF2 was differentially up-regulated after stimulation with Vibrio vulnificus, Staphylococcus aureus, Saccharomyces cerevisiae and Singapore grouper iridovirus (SGIV). Furthermore, EcILF2 was able to activate human IL-2 promoter in different cell lines and promote the endogenous IL-2 transcription in human H9 T cells. These results suggest that EcILF2 is potentially involved in grouper immune responses to invasion of bacterial and viral pathogens.
State Key Laboratory of Biocontrol, School of life sciences, Sun Yat-sen University, 135 West Xingang Road, Guangzhou 510275, PR China.
Huachun
H
Cui
Yang
Y
Yan
Jingguang
J
Wei
Zhiwei
Z
Hou
Youhua
Y
Huang
Xiaohong
X
Huang
Qiwei
Q
Qin
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004266:DNA Virus Infections / Q000276:immunology / Q000378:metabolism / Q000662:veterinary; D005393:Fish Diseases / Q000378:metabolism / Q000821:virology; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000502:physiology*; D007497:Iridoviridae; D008969:Molecular Sequence Data; D051903:Nuclear Factor 45 Protein / Q000235:genetics / Q000378:metabolism*; D010473:Perciformes / Q000378:metabolism*; D014735:Vibrio Infections
D000906:Antibodies; D051903:Nuclear Factor 45 Protein