The Concerted Conformational Changes during Human Rhinovirus 2 Uncoating

Hewat, E.A.; Neumann, Emmanuelle; Blaas, Dieter

doi:10.1016/s1097-2765(02)00603-2

Cited by 71 publications

(79 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For picornaviruses, it is supposed that after binding to the cellular receptors, an iris-type movement of major capsid protein VP1 occurs to open up a channel at the fivefold axes, which leads to exposure of the N terminus of VP1. Minor capsid protein VP4, which clusters on the inner surface under the fivefold channel, is assumed to be externalized through this channel, and the viral RNA genome is subsequently released into the cytoplasm (8,30,31). Reoviruses potentially release newly synthesized mRNAs transcribed within the intact capsid through pores at the fivefold symmetry axes (18,52,81).…”

Section: Discussionmentioning

confidence: 99%

Mutational Analysis of Narrow Pores at the Fivefold Symmetry Axes of Adeno-Associated Virus Type 2 Capsids Reveals a Dual Role in Genome Packaging and Activation of Phospholipase A2 Activity

Bleker

Sonntag

Kleinschmidt

2005

J Virol

156

185

View full text Add to dashboard Cite

Adeno-associated virus type 2 (AAV2) capsids show 12 pores at the fivefold axes of symmetry. We mutated amino acids which constitute these pores to investigate possible functions of these structures within the AAV2 life cycle. Mutants with alterations in conserved residues were impaired mainly in genome packaging or infectivity, whereas few mutants were affected in capsid assembly. The packaging phenotype was characterized by increased capsid-per-genome ratios. Analysis of capsid-associated DNA versus encapsidated DNA revealed that this observation was due to reduced and not partial DNA encapsidation. Most mutants with impaired infectivity showed a decreased capability to expose their VP1 N termini. As a consequence, the activation of phospholipase A2 (PLA2) activity, which is essential for efficient infection, was affected on intact capsids. In a few mutants, the exposure of VP1 N termini and the development of PLA2 activity were associated with enhanced capsid instability, which is obviously also deleterious for virus infection. Therefore, PLA2 activity seems to be required on intact capsids for efficient infection. In conclusion, these results suggest that the pores at the fivefold axes function not only as portals for AAV2 single-stranded DNA packaging but also as channels for presentation of the PLA2 domain on AAV2 virions during infection.The viral capsid can be considered as a multiprotein complex which not only protects the viral genome against extreme environmental influences outside the cell but also combines a number of functional elements which are needed for host cell recognition, intracellular trafficking, and disassembly in order to successfully deliver genetic information to the host cell (for reviews, see references 23, 60, and 75). These functional elements either are concomitantly transported with the virion as separate proteins or are part of the structural proteins themselves. Identifying such functional elements on viral capsids is one of the most attractive goals of viral structural research. Over the past few years, several such functional domains have been identified in parvoviruses (for a review, see reference 70).Adeno-associated virus (AAV) type 2 (AAV2) is a member of the parvovirus family. AAV2 requires coinfection with helper viruses, such as adenovirus or herpesvirus, for efficient reproduction (4, 9, 58) and has a nonenveloped icosahedral capsid. It encloses a 4.7-kb single-stranded DNA (ssDNA) genome containing two large open reading frames, rep and cap (61). The Rep proteins, encoded by the rep gene, are required for DNA replication, DNA encapsidation, and control of gene expression (36,45). The viral capsid is composed of three structural viral proteins (VPs), designated VP1, VP2, and VP3, which are encoded by the cap open reading frame and expressed from the p40 promoter. Capsids are formed with the VPs in an approximate molar ratio of 1:1:10 due to alternative splicing and an unconventional start codon for VP2. This ratio is also maintained in assembled capsids (6, 34). The ...

show abstract

Section: Discussionmentioning

confidence: 99%

Mutational Analysis of Narrow Pores at the Fivefold Symmetry Axes of Adeno-Associated Virus Type 2 Capsids Reveals a Dual Role in Genome Packaging and Activation of Phospholipase A2 Activity

Bleker

Sonntag

Kleinschmidt

2005

J Virol

156

185

View full text Add to dashboard Cite

show abstract

“…We have now solved the structure of complexes between empty capsids of HRV2 and Fab fragments of 2G2 by cryoelectron microscopy. This analysis reveals that the binding epitope of this antibody lies in the region that we have predicted to change the most upon the transition between the native virion and the empty capsid (9).…”

mentioning

confidence: 81%

“…The RNA is then released, resulting in an empty capsid with a sedimentation constant of 80S that is finally degraded in lysosomes (24). Using cryoelectron microscopy (cryo-EM) and X-ray structural data, we produced a model for the HRV2 empty capsid after RNA release (9). The capsid was seen to have expanded by 4%, with a relative movement of all capsid proteins.…”

mentioning

confidence: 99%

“…HRV2 was produced in a spinner culture of HeLa-H1 cells and purified by sucrose density gradient centrifugation, and empty capsids were obtained by incubation for 10 min at 56°C as described previously (9). The monoclonal antibody 2G2 was prepared from the tissue culture supernatant of cells grown in immunoglobulin G (IgG)-free medium by purification over a protein A column using an fast-protein liquid chromatography system, and Fab fragments were prepared and purified by following standard protocols (1).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Nonneutralizing Human Rhinovirus Serotype 2-Specific Monoclonal Antibody 2G2 Attaches to the Region That Undergoes the Most Dramatic Changes upon Release of the Viral RNA

Hewat

Blaas

2006

J Virol

Self Cite

View full text Add to dashboard Cite

The monoclonal antibody 2G2 has been used extensively for detection and quantification of structural changes of human rhinovirus serotype 2 during infection. It recognizes exclusively A and B subviral particles, not native virus. We have elucidated the basis of this selectivity by determining the footprint of 2G2. Since viral escape mutants obviously cannot be obtained, the structures of complexes between Fab fragments of 2G2 and 80S subviral B particles were determined by cryoelectron microscopy. The footprint of the antibody corresponds to the capsid region that we predicted would undergo the most dramatic changes upon RNA release.Human rhinoviruses (HRVs) belong to the picornavirus family. They are icosahedral with Tϭ1, pseudo-Tϭ3 symmetry, are about 30 nm in diameter, and are composed of 60 copies of each of the proteins VP1 through VP4 that encapsidate a single-stranded positive-sense RNA genome. Twelve of the 99 serotypes, the minor receptor group, bind members of the low-density lipoprotein receptor family for cell entry, whereas the remaining 87 serotypes use intercellular adhesion molecule 1 (ICAM-1) for infection.As shown for the major group virus HRV serotype 14 (HRV14) and for the minor group virus HRV2, receptor binding is followed by internalization via the clathrin-mediated endocytic pathway (8,25). Within the endosome, the virion undergoes coordinated structural changes preceding the RNA release into the cytosol. For major group viruses, these changes are catalyzed by the ICAM-1 receptor and possibly aided by low endosomal pH (22). In the case of minor group viruses, the low pH of the late endosomal compartment is the exclusive trigger for these modifications.HRV2 has been extensively analyzed with respect to these structural changes (9, 11, 12, 14-17, 20, 21). In the first step, the innermost capsid protein VP4 is released, giving rise to subviral particles sedimenting at about 135S, whereas the native virion sediments at 150S. This change in sedimentation behavior is accompanied by changes in antigenicity. The RNA is then released, resulting in an empty capsid with a sedimentation constant of 80S that is finally degraded in lysosomes (24). Using cryoelectron microscopy (cryo-EM) and X-ray structural data, we produced a model for the HRV2 empty capsid after RNA release (9). The capsid was seen to have expanded by 4%, with a relative movement of all capsid proteins. In particular, the viral protein VP1 around the fivefold axes make an iris type of movement to open a 10-Å-diameter channel, which allows the RNA genome to exit.The monoclonal antibody 2G2, obtained from a mouse injected with purified HRV2, proved extremely useful for the detection of the structural changes occurring upon infection. For example, it was employed to unequivocally demonstrate the low-pH dependency of HRV2 for infection; no subviral particles were formed in the presence of inhibitors of endosomal acidification, such as monensin (18) or the H ϩ -ATPasespecific drug bafilomycin A1 (2, 3, 5, 23). However, although the antibo...

show abstract

“…Characterization of the TrV capsid disassembly and genome release allowed us to propose a novel RNA-release mechanism for this virus Snijder et al, 2013) that differs from those proposed for members of the Picornaviridae family, i.e. Human Rhinovirus 2 (Garriga et al, 2012;Hewat et al, 2002) and Poliovirus (Bostina et al, 2011;Levy et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

Triatoma virus structural polyprotein expression, processing and assembly into virus-like particles

Sánchez-Eugenia

Méndez

Querido

et al. 2015

Journal of General Virology

View full text Add to dashboard Cite

In contrast to the current wealth of structural information concerning dicistrovirus particle structure, very little is known about their morphogenetic pathways. Here, we describe the expression of the two ORFs encoded by the Triatoma virus (TrV) genome. TrV, a member of the Cripavirus genus of the Dicistroviridae family, infects blood-sucking insects belonging to the Triatominae subfamily that act as vectors for the transmission of Trypanosoma cruzi, the aetiological agent of the Chagas disease. We have established a baculovirus-based model for the expression of the NS (non-structural) and P1 (structural) polyproteins. A preliminary characterization of the proteolytic processing of both polyprotein precursors has been performed using this system. We show that the proteolytic processing of the P1 polyprotein is strictly dependent upon the coexpression of the NS polyprotein, and that NS/P1 coexpression leads to the assembly of virus-like particles (VLPs) exhibiting a morphology and a protein composition akin to natural TrV empty capsids. Remarkably, the unprocessed P1 polypeptide assembles into quasispherical structures conspicuously larger than VLPs produced in NS/P1-coexpressing cells, likely representing a previously undescribed morphogenetic intermediate. This intermediate has not been found in members of the related Picornaviridae family currently used as a model for dicistrovirus studies, thus suggesting the existence of major differences in the assembly pathways of these two virus groups.

show abstract

The Concerted Conformational Changes during Human Rhinovirus 2 Uncoating

Cited by 71 publications

References 39 publications

Mutational Analysis of Narrow Pores at the Fivefold Symmetry Axes of Adeno-Associated Virus Type 2 Capsids Reveals a Dual Role in Genome Packaging and Activation of Phospholipase A2 Activity

Mutational Analysis of Narrow Pores at the Fivefold Symmetry Axes of Adeno-Associated Virus Type 2 Capsids Reveals a Dual Role in Genome Packaging and Activation of Phospholipase A2 Activity

Nonneutralizing Human Rhinovirus Serotype 2-Specific Monoclonal Antibody 2G2 Attaches to the Region That Undergoes the Most Dramatic Changes upon Release of the Viral RNA

Triatoma virus structural polyprotein expression, processing and assembly into virus-like particles

Contact Info

Product

Resources

About