Identification of 21 novel immune-type receptors in miiuy croaker and expression pattern of three typical inhibitory members

Abstract

Novel immune-type receptor (NITR) genes belong to the immunoglobulin superfamily and are encoded by clusters of multigene families. NITRs encode type I transmembrane proteins and are only found in teleosts. In the current study, total 21 NITR genes are identified from miiuy croaker (Miichthys miiuy) and named as MmNITR1 to MmNITR21. Miiuy croaker NITR genes that encoded one or two extracellular immunoglobulin (Ig) domains, a transmembrane (TM) region, an immunoreceptor tyrosine-based inhibitor motif (ITIM) in the cytoplasmic (Cyt) region. The majority of MmNITRs possess cytoplasmic ITIM that can be classified as inhibitory receptors. However, a smaller number of NITRs (MmNITR8, MmNITR15 and MmNITR16) can be classified as activating receptors by the lack of cytoplasmic ITIMs and presence of a positively charged residue within their transmembrane domain. As typical inhibitory receptors, MmNITR1, MmNITR2 and MmNITR3 have different characteristics of the structure. In MmNITR1 gene, variable (V) and intermediate (I) domains are encoded by two separate exons. In contrast to MmNITR1, MmNITR3 gene encode V and I domains in a single exon. And MmNITR2 gene is characterized by the presence of only one Ig-like (V-type) extracellular domain and lack of J or J-like motifs. Also MmNITR2 gene displays an additional exon which is 48bp long between the V domain and the TM region. Two and four potential N-link giycosylation sites (N-X-S/T) are present in the extracellular Ig domains. Real-time RT-PCR results showed that upon induction with Vibrio anguillarum, NITR gene expressions were induced by bacteria in kidney, liver and spleen. Meanwhile, NITRs are also primarily detected in different tissues. Phylogenetic analyses of NITR V domains indicate that MmNITR1 and MmNITR2 are more similar than MmNITR3.