Structural, Mechanistic, and Antigenic Characterization of the Human Astrovirus Capsid

Abstract

Human astroviruses (HAstVs) are nonenveloped, positive-sense, single-stranded RNA viruses that are a leading cause of viral gastroenteritis. HAstV particles display T=3 icosahedral symmetry formed by 180 copies of the capsid protein (CP), which undergoes proteolytic maturation to generate infectious HAstV particles. Little is known about the molecular features that govern HAstV particle assembly, maturation, infectivity, and immunogenicity. Here we report the crystal structures of the two main structural domains of the HAstV CP: the core domain at 2.60-Å resolution and the spike domain at 0.95-Å resolution. Fitting of these structures into the previously determined 25-Å-resolution electron cryomicroscopy density maps of HAstV allowed us to characterize the molecular features on the surfaces of immature and mature T=3 HAstV particles. The highly electropositive inner surface of HAstV supports a model in which interaction of the HAstV CP core with viral RNA is a driving force in T=3 HAstV particle formation. Additionally, mapping of conserved residues onto the HAstV CP core and spike domains in the context of the immature and mature HAstV particles revealed dramatic changes to the exposure of conserved residues during virus maturation. Indeed, we show that antibodies raised against mature HAstV have reactivity to both the HAstV CP core and spike domains, revealing for the first time that the CP core domain is antigenic. Together, these data provide new molecular insights into HAstV that have practical applications for the development of vaccines and antiviral therapies.IMPORTANCE Astroviruses are a leading cause of viral diarrhea in young children, immunocompromised individuals, and the elderly. Despite the prevalence of astroviruses, little is known at the molecular level about how the astrovirus particle assembles and is converted into an infectious, mature virus. In this paper, we describe the high-resolution structures of the two main astrovirus capsid proteins. Fitting these structures into previously determined low-resolution maps of astrovirus allowed us to characterize the molecular surfaces of immature and mature astroviruses. Our studies provide the first evidence that astroviruses undergo viral RNA-dependent assembly. We also provide new insight into the molecular mechanisms that lead to astrovirus maturation and infectivity. Finally, we show that both capsid proteins contribute to the adaptive immune response against astrovirus. Together, these studies will help to guide the development of vaccines and antiviral drugs targeting astrovirus.

Structure of a Human Astrovirus Capsid-Antibody Complex and Mechanistic Insights into Virus Neutralization

“…To determine if MAb PL-2 binds to the HAstV-2 CP spike (Spike2), the outermost domain on the virus surface, we first produced and purified recombinant Spike2 and observed that it elutes as a dimer by size exclusion chromatography, consistent with previous observations for Spike1 and Spike8 (26,27). To further show that Spike2 is folded correctly, we determined its structure to 1.87 Å ( Fig.…”

“…While the immature Tϭ3 HAstV particle contains 90 dimeric spikes, trypsin proteolysis leaves the mature Tϭ3 HAstV particle with only 30 dimeric spikes located on the 2-fold icosahedral axes. Our lab and others have determined the structures of the dimeric CP spikes from HAstV-1, HAstV-8, and turkey astrovirus 2 (26)(27)(28). We also reported the structure of the HAstV CP core domain that forms the icosahedral shell (26).…”

“…Our lab and others have determined the structures of the dimeric CP spikes from HAstV-1, HAstV-8, and turkey astrovirus 2 (26)(27)(28). We also reported the structure of the HAstV CP core domain that forms the icosahedral shell (26). The CP core domain contains two subdomains: a typical jelly roll ␤-barrel found in many RNA viruses and a squashed ␤-barrel (26).…”

“…We also reported the structure of the HAstV CP core domain that forms the icosahedral shell (26). The CP core domain contains two subdomains: a typical jelly roll ␤-barrel found in many RNA viruses and a squashed ␤-barrel (26). While it has been shown that mature HAstV gains entry into human cells via clathrin-mediated endocytosis (29), the identity of the host cell receptor(s) and the location of receptor-binding site(s) on the surface of the HAstV particle are unknown.…”

Structure of deformed wing virus, a major honey bee pathogen

“…A search for structures similar to the DWV P domain (22) identified a globular surface domain of a virus from the Astroviridae, an additional family of nonenveloped viruses like the picornaviruses and caliciviruses, all of which possess a single-stranded positive sense RNA genome (Table S2) (23). The domain is similar to the P domain of DWV in terms of having a core formed by β-strands that are complemented by short α-helices located at the periphery of the domain.…”