Novel Cathelicidins with Potent Antimicrobial, Biofilm Inhibitory, and Anti-inflammatory Activities from the Frog Fejervarya multistriata

Antimicrobial peptides (AMPs), a class of gene-encoded peptides, are the first line of immune system to defense microbial invasions in multicellular organisms. Cathelicidins are an important family of AMPs that have been identified exclusively in vertebrates. However, up to now, cathelicidins from amphibians are poorly understood. In the present study, we reported the identification and characterization of two novel cathelicidins (FM-CATH1 and FM-CATH2) from the frog Fejervarya multistriata. The cDNA sequences encoding FM-CATHs were successfully cloned from the constructed lung cDNA library of F. multistriata. Both of the cDNA sequences encoding FM-CATHs are 447 bp in length, and the deduced mature peptides of FM-CATHs are composed of 34 residues. Structural analysis indicated that FM-CATH1 and FM-CATH2 mainly assume amphipathic alpha-helical conformations. Antimicrobial and bacterial killing kinetic analysis indicated that both FM-CATH1 and FM-CATH2 possess potent, broad-spectrum and rapid antimicrobial potency. And cytoplasmic membrane permeabilization analysis indicated that FM-CATH1 and FM-CATH2 kill bacteria by inducing the permeabilization of bacterial membrane. Besides direct antimicrobial activities, FM-CATHs also exhibited significant inhibitory effect on the formation of bacterial biofilms at low concentrations below 1xMIC. Furthermore, FM-CATH1 and FM-CATH2 exhibited potent anti-inflammatory activities by inhibiting LPS-induced transcription and production of pro-inflammatory cytokines TNF-alpha, IL-1 beta, and IL-6 in mouse peritoneal macrophages. Meanwhile, FM-CATHs showed relatively low cytotoxic activity against mammalian normal and tumor cell lines, and low hemolytic activity against human erythrocytes. In summary, the identification of FM-CATHs provides novel clues for our understanding of the roles of cathelicidins in amphibian immune systems. The potent antimicrobial, biofilm inhibitory, anti-inflammatory activities, and low cytotoxicity of FM-CATHs imply their great potential in novel antibiotics development.