Molecular cloning and characterization of IKKε gene from black carp Mylopharyngodon piceus

IKK epsilon is an I kappa B kinase functioning in NF-kappa B signal pathway in the innate immune system of higher vertebrates. To exploit the function of IKK epsilon of black carp (bcIKK epsilon) in its antiviral innate immunity, the IKK epsilon gene has been cloned from the RNA isolated from the spleen of black carp. The full-length cDNA of bcIKK epsilon is 2537 bp, which encodes the peptide of 723 amino acids. bcIKK epsilon contains a S-Tkc domain, a PKc domain and a UBL-TBK1-like domain and bcIKK epsilon shares the highest amino acid sequence similarity with that of grass carp. bcIKK epsilon was constitutively transcribed in the selected tissues of black carp including gill, kidney, heart, intestine, liver, muscle, skin and spleen; and the mRNA level of bcIKK epsilon in these tissues varied right after SVCV or GCRV infection. bcIKK epsilon had been well expressed in HEK293T cells and western blot assay determined that this fish kinase was around 80 KDa. The immunofluorescence assay of both NH3T3 cells and EPC cells demonstrated that bcIKK epsilon was located in the cytosolic part of the cell. Report assay result showed that overexpression of bcIKK epsilon in EPC cells activated the expression of both zebrafish IFN and EPC IFN. All our data suggest that bcIKK epsilon is a novel fish kinase functioning in the innate antiviral immune response of black carp. (C) 2015 Elsevier Ltd. All rights reserved.