Human alpha-defensin 5 stabilizes the enterovirus A71 capsid and blocks infection

Abstract

Human alpha-defensins are antimicrobial peptides abundantly expressed in neutrophils and the small intestine. They block infection of several families of non-enveloped DNA viruses by binding to and stabilizing the viral capsid during entry, thereby preventing the genome from reaching the nucleus to initiate replication. It is unclear if a similar mechanism also applies to RNA viruses. To study this further, we investigated the interaction of human alpha-defensin 5 (HD5) with enterovirus A71 (EV-A71). We found that HD5 disrupts EV-A71 infection in cell culture and blocks viral entry. HD5 binds directly to the EV-A71 capsid and disrupts key conformational changes essential to the initiation of in vitro uncoating as well as downstream viral genome release. Using a suite of HD5 point mutants, we found that these two uncoating blocks are separable, and HD5 must achieve both to fully neutralize EV-A71 infection. This work advances our understanding of alpha-defensin antiviral action and demonstrates several conserved features of HD5 inhibition that expand to a clinically important RNA virus. Author SummaryIn this study, we demonstrated that human defensin 5 (HD5) inhibits infection of pathogenic enterovirus A71 (EV-A71). HD5 binds to EV-A71 and disrupts in vitro viral uncoating by stabilizing the viral capsid and blocking genome release. Using a library of HD5 mutants, we identified a continuous surface within HD5 important for binding to EV-A71 and disrupting infection. This surface includes amino acid residues previously shown to be important for inhibition of human adenovirus 5 and human papillomavirus 16 and represents a conserved interface involved in broad-spectrum HD5 antiviral activity. We determined that capsid stabilization and blocked genome release are independent modes of HD5 action and identified individual HD5 mutants with separable functions. These mutants provide valuable tools to further study the biology of EV-A71 uncoating. Overall, this work demonstrates the first example of an enterovirus inhibited by an -defensin and advances our mechanistic understanding of HD5 antiviral action to a new pathogenic virus.