A Novel Open and Infectious Form of Echovirus 1

Abstract

One of the hallmarks of enterovirus genome delivery is the formation of an uncoating intermediate particle. Based on previous studies of mostly heated picornavirus particles, intermediate particles were shown to have externalized the innermost capsid protein (VP4) and exposed the N terminus of VP1 and to have reduced infectivity. Here, in addition to the native and intact particle type, we have identified another type of infectious echovirus 1 (E1) particle population during infection. Our results show that E1 is slightly altered during entry, which leads to the broadening of the major virion peak in the sucrose gradient. In contrast, CsCl gradient separation revealed that in addition to the light intact and empty particles, a dense particle peak appeared during infection in cells. When the broad peak from the sucrose gradient was subjected to a CsCl gradient, it revealed light and dense particles, further suggesting that the shoulder represents the dense particle. The dense particle was permeable to SYBR green II, it still contained most of its VP4, and it was able to bind to its receptor ␣ 2 ␤ 1 integrin and showed high infectivity. A thermal assay further showed that the ␣ 2 ␤ 1 integrin binding domain (I-domain) stabilized the virus particle. Finally, heating E1 particles to superphysiological temperatures produced more fragile particles with aberrant ultrastructural appearances, suggesting that they are distinct from the dense E1 particles. These results describe a more open and highly infectious E1 particle that is naturally produced during infection and may represent a novel form of an uncoating intermediate. IMPORTANCE In this paper, we have characterized a possible uncoating intermediate particle of E1 that is produced in cells during infection.Before releasing their genome into the host cytosol, enteroviruses go through structural changes in their capsid, forming an uncoating intermediate particle. It was shown previously that structural changes can be induced by receptor interactions and, in addition, by heating the native virion to superphysiological temperatures. Here, we demonstrate that an altered, still infectious E1 particle is found during infection. This particle has a more open structure, and it cannot be formed by heating. It still contains the VP4 protein and is able to bind to its receptor and cause infection. Moreover, we show that in contrast to some other enteroviruses, the receptor-virion interaction has a stabilizing effect on E1. This paper highlights the differences between enterovirus species and further increases our understanding of various uncoating forms of enteroviruses. Picornaviruses are a large family of pathogens infecting humans and animals across the globe. The vast disease range caused by these viruses spans from simple rashes to paralysis and meningitis. It has been estimated that approximately half of the seasonal common-cold cases reported are caused by rhinoviruses, members of the family subgroup Enterovirus (1). Increased emphasis on enterovirus research is...

Extracellular Albumin and Endosomal Ions Prime Enterovirus Particles for Uncoating That Can Be Prevented by Fatty Acid Saturation

“…7). Moreover, the treated virus sample showed approximately 2 logs lower infectivity (decrease from 8.23 ϫ 10 11 to 1.01 ϫ 10 10 ), confirming our previously published data for the E1 uncoating intermediate particle (14). Hence, VP4 could contribute to the poorly ordered density on the inside of the capsid close to the vertices, attributed primarily to RNA, as well as to the density spanning the capsid.…”

“…We showed previously that during entry into cells, E1 undergoes structural changes that were first discovered as an increased permeability to the small molecule dye, SYBR green II, and Cs-ions (14). Here, we showed by cryoEM that temperature-dependent structural changes, under physiological conditions, involved expansion of the virus particle, loosening of the genome packing, loss of the lipid factor, and formation of larger openings at the VP2 dimer interface, explaining the increased permeability to the small molecular dye and the resistance to RNase treatment observed previously (14). We also showed by spectroscopy that serum priming of coxsackievirus B3 and coxsackievirus A9 caused similar changes in permeability to that of E1, suggesting that also other enteroviruses behave similarly (Fig.…”

“…Recently, we described a novel uncoating intermediate particle of E1 which formed during infection and could be isolated from the cells at early stages of infection (14). This particle proved to be stable, infectious, containing all of its capsid proteins, and still capable of receptor binding (14).…”

“…Recently, we described a novel uncoating intermediate particle of E1 which formed during infection and could be isolated from the cells at early stages of infection (14). This particle proved to be stable, infectious, containing all of its capsid proteins, and still capable of receptor binding (14). Previously, several studies characterized uncoating intermediates for entero-and rhinoviruses that have been termed as A-or 135Sparticles based on their altered conformation and lighter sedimentation in sucrose gradients (15).…”

Novel activities of safe-in-human broad-spectrum antiviral agents

“…Human influenza A/WSN/33(H1N1) virus was generated using eight-plasmid reverse genetics system in HEK293 and Vero-E6 cells, as described previously (Hoffmann et al, 2000). EV1 strain was propagated in a monolayer of Vero cells, as described earlier (Myllynen et al, 2016). HSV-1 was amplified in Vero cells, as described previously (Nygardas et al, 2013).…”