Cloning and expression analysis of interferon induced protein 44 gene in Nile tilapia(Oreochromis niloticus)

【Objective】 The expression characteristics among different tissues and the expression pattern of interferon induced protein 44 gene(ifi44) after Streptococcus agalactiae infection and polyinosinic-polycytidylic acidPoly(I:C) challenge were explored in present study to lay a theoretical basis for further researching the function and mechanism by ifi44 in the immune response of fish.【Method】 The open reading frame(ORF) of On-ifi44 gene was cloned and its bioinformatics was analyzed by ExPASy, TMHMM-2.0, Euk-mPLoc 2.0, SOPMA and SWISS-MODEL online softwares. The single-cell RNA-Seq(scRNA-seq) data were employed to analyze the distribution of On-ifi44 gene at the cellular level. The expression distribution of On-ifi44 gene in various tissues of healthy Nile tilapia and the sequential expression after S. agalactiae infection and Poly(I:C) stimulation were detected by real-time fluorescent quantitative PCR.【Result】 The open reading frame of the On-ifi44 gene was 1437 bp, encoding 478 amino acids residues with a protein molecular weight of 52.7 kD and theoretical isoelectric point(pI) of 4.97. On-ifi44 contained a TLDc_dom domain(1-157 aa) and a GTP-bd domain(197-322 aa), but no transmembrane domain. Amino acid sequence similarity between On-ifi44 and Oreochromis aureus ifi44 was 97.94%, phylogenetic tree based on amino acid sequence similarity of ifi44 amino acid sequence similarity showed that On-ifi44 amino acid sequence and O. aureus ifi44 amino acid sequence clustered into a single lineage, which meaned that they had a close genetic relationship. On-ifi44 gene was expressed in 11 tissues of Nile tilapia, including intestine, liver, gills, skin, spleen, kidney, thymus, brain, head-kidney, muscle and heart, and mainly expressed in the T cell subgroup. After S. agalactiae infection and Poly(I:C) stimulation, the expression of On-ifi44 gene was changed significantly in the spleen, kidney, head-kidney, and intestinal, and with obvious time-dependence.【Conclusion】 On-ifi44 contains one TLDc_dom domain and one GTP-bd domain, which is evolutionarily conserved and has similar biological functions among different species. The expression of On-ifi44 gene is up-regulated in a time-dependent manner after S. agalactiae infection and Poly(I:C) stimulation, indicating that ifi44 gene, as an important regulatory factor, is involved in the immune response of Nile tilapia against bacterial infection and viral proliferation.