A defensin from clam Venerupis philippinarum: Molecular characterization, localization, antibacterial activity, and mechanism of action

Antimicrobial peptides (AMPs) are important mediators of the primary host defense system against microbial invasion. In the present study, we cloned and characterized a member of the invertebrate defensin from the clam Venerupis philippinarum, designated VpDef. Amino acid sequence analysis showed that VpDef was similar to defensins from marine mollusks and ticks. In non-stimulated clams, RT-PCR and immunohistochemical analysis revealed that both VpDef mRNA and the encoding peptide were constitutively expressed in hemocytes and mantles, as well as in other major tissues. VpDef transcripts were significantly induced in hemocytes at different time intervals post Vibrio anguillarum infection. The recombinant VpDef (rVpDef) showed the highest activity against Gram-positive bacteria Micrococcus luteus and less effective to Gram-negative bacteria. In addition, incubation of rVpDef with M. luteus at 1 × and 3 × MIC could induce an obvious decrease of the membrane potential and notable changes of membrane permeability in a dose-dependent manner. Membrane integrity and bacterial viability analysis also revealed that rVpDef increased the membrane permeability of M. luteus and then resulted in cell death at 2 × and 10 × MIC. Overall, these results suggest that VpDef has an important function in host defense against invasive pathogens, probably killing microbes by inducing membrane lesions.