Cloning and expression analysis of IKKβ gene in Penaeus japonicus

【Objective】 To elucidate the biological characteristics of NF-κB inhibitory protein kinase-β gene（named PjIKKβ）in Penaeus japonicus and its expression pattern in various tissues and stimulated by Vibrio harveyi ，and to provide theoretical basis for revealing the mechanism of IKKβ in the innate immunity of P. japonicus.【Method】The cDNA sequence of PjIKKβ gene was cloned. The bioinformatics of PjIKKβ gene was analyzed by some online softwares，such as ORF Finder，ExPASy ProtParam，SMART，SignalP 5.0，TMHMM and Prot-Comp. The expression of PjIKKβ gene was examinated in various tissues under normal condition and in haemolymph after the treatment with V. harveyi by real-time fluorescence quantitative PCR.【Result】 The full length of PjIKKβ gene sequence was 2794 bp and the open reading frame （ORF）was 2373 bp which encoding 790 amino acid residues. PjIKKβ protein contained a serine/threonine protein kinases catalytic（S-TKc）domain，a leucine zipper（LZ）domain and a helix loop helix（HLH）domain. PjIKKβ protein was localized in cytoplasm and nucleus. No transmembrane structure and signal peptide were detected in PjIKKβ protein. PjKKβ protein shared the highest identity with IKKβ amino acid sequence of P. vannamei（98%）and P. monodon（97%）. Multiple sequence alignment analysis showed that S-TKc domain was highly conserved among various species and other domains were low conservation. The result of phylogenetic tree constructed based on IKKβ amino acid sequence showed that P. japonicus were clustered firstly with P. vannamei and then clustered with P. vannamei. PjIKKβ gene was ubiquitously expressed in all tested tissues（hepatopancreas，muscles，gills，ganglia，stomach，heart，hemolymph and eye stalk）. The highest expression level was detected in the ganglion，muscle，haemolymph and eye stalk. The expression of PjIKKβ gene in haemolymph was up-regulated at 24，48 and 72 h after infected of V. harveyi and was extremely significantly different from that infected with PBS（P＜0.01）.【Conclusion】 PjIKKβ protein contains a S-TKc domain，a LZ domain and a HLH domain，which is conservative in evolution. The expression of PjIKKβ in various tissues and its changes of expression after stimulation by V. harveyi indicate that PjIKKβ is involved in the innate immune response of P. japonicus.