Expression analysis and SNP development of the NF-κB1 gene in Siniperca chuatsi against Aeromonas hydrophila

【Objective】 This study aimed to explore the function of nuclear factor kappa B subunit 1 （NF-κB1） gene in the immune response in Siniperca chuatsi against Aeromonas hydrophila， and to explore the related single nucleotide polymorphism （SNP） loci， so as to provide theoretical basis for revealing the molecular mechanism of bacterial infection resistance and disease resistance breeding of Siniperca chuatsi.【Method】 Bioinformatics analysis was carried out to analyze characteristics of NF-κB1 gene sequences in Siniperca chuatsi. Real-time fluorescence quantitative PCR was used to detect the relative expressions of NF-κB1 gene in various tissues before and after Aeromonas hydrophila infection. The regulation of NF-κB1 gene on pro-inflammatory genes IL-1β and TNF-α was detected by siRNA interference. Combined with Sanger sequencing and association analysis， SNP loci significantly associated with disease resistance traits in NF-κB1 gene were screened.【Result】 The length of the coding region of NF-κB1 gene of Siniperca chuatsi was 2553 bp， encoding 850 amino acid residues， including four domains of RHD-n， IPT， ANKYR and Death. Among them， RHD-n and IPT domains were the most conserved in evolution. The NF-κB1 protein in Siniperca chuatsi was clustered with that in fishes such as Epinephelus lanceolatus， with a high similarity of amino acid sequences. The relative expression of NF-κB1 gene in muscle tissue was significantly higher than that in head kidney， spleen， and other tissues （P<0.05， the same below）. After Aeromonas hydrophila infection， the relative expressions of the NF-κB1 gene in liver， head kidney， and muscle tissues increased. After siRNA interference of NF-κB1， the relative expressions of IL-1β and TNF-α genes in muscle was significantly down-regulated. The SNP3 locus （g.1025140 C>A） in the intron region of NF-κB1 gene was significantly associated with the resistance of Siniperca chuatsi against Aeromonas hydrophila.【Conclusion】 The NF-κB1 gene of Siniperca chuatsi plays a key role in the core immune pathway against bacterial infection through its highly conserved RHD-n domain. The NF-κB1 gene positively regulates the expression of key pro-inflammatory genes such as IL-1β and TNF-α after infection through typical signaling pathways， thereby driving the early inflammatory defense response of Siniperca chuatsi. At the same time， the SNP3 locus （g.1025140 C>A） in the intron region of the NF-κB1 gene is significantly associated with disease resistance， indicating important regulatory elements may appear in this region.