Molecular Cloning of Cathelicidin-like cDNA from Andrias davidianus

Abstract

Antimicrobial peptides (AMPs) as part of host defense systems has been widely recognized in most organisms. Cathelicidin is an important family of AMPs acting as multifunctional effector molecules in innate immunity and exists in organisms with cathelicidin-like precursor. Andrias davidianus (A. davidianus) is a unique species in China and the biggest amphibians in the world. With the rapid growth of A. davidianus aquaculture, pathogens of bacteria, virus and fungus were reported, however little is known about antimicrobial peptides derived from A. davidianus. To investigate antimicrobial peptides of cathelicidin-like in A. davidianus, cathelicidin-like precursor gene cloning and bioinformatic analysis was carried out. The results showed that 1106 bp full-length cDNA of cathelicidin-like precursor was obtained, which was including a 35 bp 5' terminal UTR, a 546 bp open reading frame (ORF) and a 525 bp 5' terminal UTR. The cathelicidin-like precursor amino acid (AA) sequence of A. davidianus comprised N-terminal signal peptide (21 AA), highly conserved cathelin domain and C-terminal mature peptide. The cathelicidin-like precursor gene nucleotide sequence showed low identify with other cathelicidin-like sequences, while AA sequence displayed relatively higher similarity with cathelicidin-like isolated from other species. Phylogenetic tree indicated cathelicidinlike precursor of A. davidianus was firstly clade with Tylototrition verrucosus, which also belonged to Caudata, Amphibian. The precursor gene expression was detected by RT-qPCR. The result displayed this gene was abundant expression in A. davidianus skin. According the specificity proteases cleavage and characteristic of cathelicidin, five putative mature cathelicidin were predicted. This study confirms the presence of cathelicidin in A. davidianus. Their results not only reveal innate immune system of A. davidianus but also enlarge the AMP knowledge of urodele amphibians.