MOLECULAR IDENTIFICATION AND INNATE IMMUNE ROLE OF FIRST TELEOST MALECTIN FROM Hippocampus abdominalis
In this study, we have identified and characterized ER resident lectin type protein, malectin from big-belly seahorse Hippocampus abdominalis (HaMLEC). Lectins are carbohydrate-binding proteins have a role in cellular and molecular level recognition and also mediate bacterial and viral binding to the targets. Malectin is involving in backup glycol-protein quality control in endoplasmic reticulum (ER) and have binding specificity to Glc2-N-glycan. Also, malectin contains CBMs like structure, which might have the carbohydrate and bacterial binding ability. Malectin indicated to have a potential role in innate immunity during some recent studies. However, the studies on innate immune role of malectin is very limited.
The ORF and protein sequence of HaMLEC was deduced by UGENE software and the in-silico study was performed with the deduced protein sequence of HaMLEC by using bioinformatic tools. We have analysed the spatial and temporal transcript level of HaMLEC by qPCR detection. Immune challenge was carried out with four immune stimulants (Edwardasiella tarda, Streptococcus iniae, polyinosinic: polycytidylic, and lipopolysaccharide) and the liver sample was collected at different time points for the temporal level analysis. The bacterial binding ability of HaMLEC was analysed through ELISA by using recombinant HaMLEC with different bacteria (E. tarda, Lactococcus garvieae, V. anguillarum, S. iniae, Vibrio harveyi, S. parauberis, and E. coli).
The cDNA sequence of HaMLEC consists of 1269 bp ORF, which encoding 423 amino acids. The molecular weight was predicted as 49.04 kDa and the pI was estimated as 4.37. Malectin super family, C-terminal transmembrane region, and N-terminal signal peptide was identified from HaMLEC sequence. The HaMLEC showed high sequence identity (99.0 %) and close evolutionary relationship with Hippocampus comes during pairwise sequence alignment and phylogenetic analysis, respectively. The transcript level of HaMLEC was detected in all tested tissues. The significant upregulations were observed for all four stimulants with different time points during temporal level expression in liver. Moreover, the significant bacterial binding ability was observed with wide range of bacterial species with compare to the controls. The observed significant changes in immune challenge results and the wide range of bacterial binding activity suggesting that the HaMLEC might be involved in the innate immune role of Hippocampus abdominalis.
