Molecular characterization, expression and functional analysis of a nuclear oligomerization domain proteins subfamily C (NLRC) in Japanese flounder (Paralichthys olivaceus)

Pattern recognition receptors (PRRs) are involved in the effective innate defense against several microbes. Here, we identified a nucleotide-oligomerization domain (NOD)-like receptor subfamily C (NLRC) from Japanese flounder (Paralichthys olivaceus). Full-length transcript of JfNLRC is composed of 3976 bp encoding a protein of 1175 deduced amino acid residues. The presence of a signature nucleotide-binding domain (NACHT) and leucine-rich repeated domain (LRR) suggested that the protein is a member of the NLR family. Interestingly, its C-terminus presents an extra PRY/SPRY (B30.2) domain similar to fish in the Trim (finTrim) family. A phylogenic tree of JfNLRC revealed that full-length JfNLRC diverged from the NOD1 and NOD2 clusters, and the NACHT domain in JfNLRC was clustered within the NLRC3 group. Stimulation by formalin-killed Edwardsiella tarda, Streptococcus iniae, and lipopolysaccharide (LPS) showed that the JfNLRC expression was raised a few hours after stimulation, suggesting this novel protein is involved in the immediate response against both Gram-positive and Gram-negative bacteria. Furthermore, the IL-1β mRNA expression level in JfNLRC-over-expressing HINAE cells was significantly increased, when compared to a control, after LPS-stimulation and E. tarda infection. These results suggested that JfNLRC probably induced IL-1β gene expression mediated by LPS-stimulation.

► NLRC mRNA in Japanese flounder was cloned.
► The phylogenetic tree was analyzed and evolutionary conservation was clarified.
► The tissue distribution of NLRC was measured.
► NLRC transcripts were increased by stimulation with formalin-killed bacteria and LPS.
► NLRC probably mediates induction of IL-1b mRNA expression through LPS-stimulation.