The Structure of Echovirus Type 12 Bound to a Two-domain Fragment of Its Cellular Attachment Protein Decay-accelerating Factor (CD 55)

Decay-accelerating factor (DAF) functions as cell attachment receptor for a wide range of human enteroviruses. The Kuykendall prototype strain of coxsackievirus A21 (CVA21) attaches to DAF but requires interactions with intercellular cell adhesion molecule 1 (ICAM-1) to infect cells. We show here that a bioselected variant of CVA21 (CVA21-DAFv) generated by multiple passages in DAF-expressing, ICAM-1-negative rhabdomyosarcoma (RD) cells acquired the capacity to induce rapid and complete lysis of ICAM-1-deficient cells while retaining the capacity to bind ICAM-1. CVA21-DAFv binding to DAF on RD cells mediated lytic infection and was inhibited by either antibody blockade with a specific anti-DAF SCR1 monoclonal antibody (MAb) or soluble human DAF. Despite being bioselected in RD cells, CVA21-DAFv was able to lytically infect an additional ICAM-1-negative cancer cell line via DAF interactions alone. The finding that radiolabeled CVA21-DAFv virions are less readily eluted from surface-expressed DAF than are parental CVA21 virions during a competitive epitope challenge by an anti-DAF SCR1 MAb suggests that interactions between CVA21-DAFv and DAF are of higher affinity than those of the parental strain. Nucleotide sequence analysis of the capsid-coding region of the CVA21-DAFv revealed the presence of two amino acid substitutions in capsid protein VP3 (R96H and E101A), possibly conferring the enhanced DAF-binding phenotype of CVA21-DAFv. These residues are predicted to be embedded at the interface of VP1, VP2, and VP3 and are postulated to enhance the affinity of DAF interaction occurring outside the capsid canyon. Taken together, the data clearly demonstrate an enhanced DAF-using phenotype and expanded receptor utilization of CVA21-DAFv compared to the parental strain, further highlighting that capsid interactions with DAF alone facilitate rapid multicycle lytic cell infection.The attachment of viruses to cell surface molecules is the initial step of virus replication, and specific cellular virus receptors are therefore major determinants for virus tissue tropism. Decay-accelerating factor (DAF; CD55), a 70-kDa glycosylphosphatidylinositol-anchored complement-regulatory protein consisting of four extracellular short consensus repeats (SCRs) (25), serves as a membrane attachment protein for numerous human enteroviruses, including several echoviruses (EV) (4,20,36,48), coxsackie B viruses (CVB) (41) and coxsackievirus A21 (CVA21) (43). In general, viral binding to DAF alone is insufficient to permit enteroviral infections and interactions with DAF do not induce 135S altered (A) particles (34,35,39,43,45), which are considered to be a prerequisite for cell entry (2, 50). The physiological role of DAF for enteroviral infections is postulated to be that of a membrane concentration receptor that binds and clusters the infectious virus, resulting in increased opportunity for cell entry via interactions with a second functional cell entry receptor (30,43,45).As with many other picornaviruses (the polioviruses, the major-rece...

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Sedimentation Of Daf-and Icam-1-bound Virionsmentioning

confidence: 99%

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Enhanced Cellular Receptor Usage by a Bioselected Variant of Coxsackievirus A21

Johansson

Cheng

et al. 2004

“…Receptor interaction with the canyon may lead to displacement of the pocket factor and destabilization of the capsid (34). Although the binding site for DAF on the CB3 capsid has not been directly identified, genetic evidence (26,30) and the structures of complexes between DAF and other enteroviruses (5,19) suggest that DAF binds outside the canyon, where it is unlikely to trigger capsid disruption.…”

Section: A-particle Formation Is Induced By Car But Not By Daf Onmentioning

confidence: 99%

Interaction with Coxsackievirus and Adenovirus Receptor, but Not with Decay-Accelerating Factor (DAF), Induces A-Particle Formation in a DAF-Binding Coxsackievirus B3 Isolate

Milstone

Petrella

Sánchez

et al. 2005

Although many coxsackie B viruses interact with decay accelerating factor (DAF), attachment to DAF by itself is not sufficient to initiate infection. We examined the early events in infection that follow virus interaction with DAF, and with the coxsackievirus and adenovirus receptor (CAR). Interaction with soluble CAR in a cell-free system, or with CAR on the surfaces of transfected cells, induced the formation of A particles; interaction with soluble or cell surface DAF did not. The results suggest that CAR, but not DAF, is capable of initiating the conformational changes in the viral capsid that lead to release of viral nucleic acid.

“…Direct involvement of ␣ v integrins in the infectious entry of HPEV1 was further confirmed by overexpression of human ␣ v ␤ 1 and ␣ v ␤ 3 integrins in Chinese hamster ovary (CHO) cells, allowing successful virus infection (74). There are no reports yet on the identification of receptors for the HPEV types lacking the RGD motif (HPEV3, HPEV7, and HPEV8) (19,39,51).Although the crystal structures of several picornaviruses have been determined (3,26,34,35,44,57,59,65,68,72) and the receptor interactions have been studied in detail by X-ray crystallography, electron cryo-microscopy (cryo-EM), and threedimensional (3D) image reconstruction (6,9,23,31,32,47,83), there is no structural information available for the parechoviruses or parechovirus-receptor complexes. Here, we compare the binding of ␣ V ␤ 3 and ␣ V ␤ 6 to HPEV1 in vitro by biochemical assays and determine the structures of HPEV1 and the corresponding HPEV1-integrin complexes.…”

mentioning

confidence: 99%

Interaction of α _V β ₃ and α _V β ₆ Integrins with Human Parechovirus 1

Seitsonen

Susi

Heikkilä

et al. 2010

Human parechovirus (HPEV) infections are very common in early childhood and can be severe in neonates. It has been shown that integrins are important for cellular infectivity of HPEV1 through experiments using peptide blocking assays and function-blocking antibodies to ␣ V integrins. The interaction of HPEV1 with ␣ V integrins is presumably mediated by a C-terminal RGD motif in the capsid protein VP1. We characterized the binding of integrins ␣ V ␤ 3 and ␣ V ␤ 6 to HPEV1 by biochemical and structural studies. We showed that although HPEV1 bound efficiently to immobilized integrins, ␣ V ␤ 6 bound more efficiently than ␣ V ␤ 3 to immobilized HPEV1. Moreover, soluble ␣ V ␤ 6, but not ␣ V ␤ 3, blocked HPEV1 cellular infectivity, indicating that it is a high-affinity receptor for HPEV1. We also showed that HPEV1 binding to integrins in vitro could be partially blocked by RGD peptides. Using electron cryo-microscopy and image reconstruction, we showed that HPEV1 has the typical T‫1؍‬ (pseudo T‫)3؍‬ organization of a picornavirus. Complexes of HPEV1 and integrins indicated that both integrin footprints reside between the 5-fold and 3-fold symmetry axes. This result does not match the RGD position predicted from the coxsackievirus A9 X-ray structure but is consistent with the predicted location of this motif in the shorter C terminus found in HPEV1. This first structural characterization of a parechovirus indicates that the differences in receptor binding are due to the amino acid differences in the integrins rather than to significantly different viral footprints.Picornaviruses consist of a positive-sense, single-stranded infectious RNA genome of approximately 7.3 kb enclosed in a capsid composed of 60 copies of each of the three or four capsid proteins (VP1 to VP4). Human parechovirus 1 (HPEV1) is a member of the Parechovirus genus of the Picornaviridae family (38, 70). There are currently eight completely sequenced human parechovirus types and 14 described types (4,19,24,30,38,39,51,58,78). In addition, the Parechovirus genus currently has four Ljungan virus members that infect rodents. HPEV1 exhibits several distinct molecular characteristics compared to other picornaviruses (38, 71). These include the lack of the maturation cleavage of the capsid proteins VP0 to VP4 (Nterminal) and VP2 (C-terminal), existence of an approximately 30-amino-acid-long extension to the N terminus of VP3, a unique nonstructural protein 2A, and a 5Ј untranslated region that is more closely related to picornaviruses infecting animals than those infecting humans.HPEV infections are common during the first years of life and are often mild or asymptomatic (20,28,42,73,80). Recently, a number of new types have been identified, and their prevalence in stool samples, for example, highlights their clinical importance. Normally, they cause gastroenteritis and respiratory infections, but severe illnesses, such as infections of the central nervous system, generalized infections of neonates, and myocarditis, have also been associated with HPEV infect...