Parvovirus particles as platforms for protein presentation.

Miyamura, Koichi; Kajigaya, Sachiko; Momoeda, Mikio; Smith‐Gill, Sandra J.; Young, Neal S.

doi:10.1073/pnas.91.18.8507

Cited by 39 publications

(20 citation statements)

References 34 publications

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“…Baculovirus-expressed B19 capsids containing truncated Flag-VP1 were recognized by an anti-Flag monoclonal antibody (MAb) (19). Similarly, baculovirus-derived B19 capsids in which VP1u was replaced with lysozyme were enzymatically active and immunogenic (26). These results suggest that VP1u occupies an external position on the capsid.…”

mentioning

confidence: 67%

Conformational Changes in the VP1-Unique Region of Native Human Parvovirus B19 Lead to Exposure of Internal Sequences That Play a Role in Virus Neutralization and Infectivity

Ros

Gerber

Kempf

2006

J Virol

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The unique region of the capsid protein VP1 (VP1u) of human parvovirus B19 (B19) elicits a dominant immune response and has a phospholipase A 2 (PLA 2 ) activity, which is necessary for the infection. In contrast to the rest of the parvoviruses, the VP1u of B19 is thought to occupy an external position in the virion, making this region a promising candidate for vaccine development. By using a monoclonal antibody against the most-N-terminal portion of VP1u, we revealed that this region rich in neutralizing epitopes is not accessible in native capsids. However, exposure of capsids to increasing temperatures or low pH led to its progressive accessibility without particle disassembly. Although unable to bind free virus or to block virus attachment to the cell, the anti-VP1u antibody was neutralizing, suggesting that the exposure of the epitope and the subsequent virus neutralization occur only after receptor attachment. The measurement of the VP1u-associated PLA 2 activity of B19 capsids revealed that this region is also internal but becomes exposed in heat-and in low-pH-treated particles. In sharp contrast to native virions, the VP1u of baculovirus-derived B19 capsids was readily accessible in the absence of any treatment. These results indicate that stretches of VP1u of native B19 capsids harboring neutralizing epitopes and essential functional motifs are not external to the capsid. However, a conformational change renders these regions accessible and triggers the PLA 2 potential of the virus. The results also emphasize major differences in the VP1u conformation between natural and recombinant particles.

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mentioning

confidence: 67%

Conformational Changes in the VP1-Unique Region of Native Human Parvovirus B19 Lead to Exposure of Internal Sequences That Play a Role in Virus Neutralization and Infectivity

Ros

Gerber

Kempf

2006

J Virol

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“…These data open up the realistic possibility of using the BTV CLP particle as a delivery system for epitopes or enzymes to targeted cells (12,26,38).…”

Section: Discussionmentioning

confidence: 99%

Assembly and Intracellular Localization of the Bluetongue Virus Core Protein VP3

Kar

Iwatani

Roy

2005

J Virol

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The bluetongue virus (BTV) core protein VP3 plays a crucial role in the virion assembly and replication process. Although the structure of the protein is well characterized, much less is known about the intracellular processing and localization of the protein in the infected host cell. In BTV-infected cells, newly synthesized viral core particles accumulate in specific locations within the host cell in structures known as virus inclusion bodies (VIBs), which are composed predominantly of the nonstructural protein NS2. However, core protein location in the absence of VIBs remains unclear. In this study, we examined VP3 location and degradation both in the absence of any other viral protein and in the presence of NS2 or the VP3 natural associate protein, VP7. To enable real-time tracking and processing of VP3 within the host cell, a fully functional enhanced green fluorescent protein (EGFP)-VP3 chimera was synthesized, and distribution of the fusion protein was monitored in different cell types using specific markers and inhibitors. In the absence of other BTV proteins, EGFP-VP3 exhibited distinct cytoplasmic focus formation. Further evidence suggested that EGFP-VP3 was targeted to the proteasome of the host cells but was dispersed throughout the cytoplasm when MG132, a specific proteasome inhibitor, was added. However, the distribution of the chimeric EGFP-VP3 protein was altered dramatically when the protein was expressed in the presence of the BTV core protein VP7, a normal partner of VP3 during BTV assembly. Interaction of EGFP-VP3 and VP7 and subsequent assembly of core-like particles was further examined by visualizing fluorescent particles and was confirmed by biochemical analysis and by electron microscopy. These data indicated the correct assembly of EGFP-VP3 subcores, suggesting that core formation could be monitored in real time. When EGFP-VP3 was expressed in BTV-infected BSR cells, the protein was not associated with proteasomes but instead was distributed within the BTV inclusion bodies, where it colocalized with NS2. These findings expand our knowledge about VP3 localization and its fate within the host cell and illustrate the assembly capability of a VP3 molecule with a large amino-terminal extension. This also opens up the possibility of application as a delivery system. Bluetongue virus (BTV) is anOrbivirus that has a nonenveloped icosahedral particle that contains an RNA genome of 10 double-stranded segments within three concentric protein shells. The outer shell is composed of two proteins, VP2 and VP5, and the inner shell, or core, consists of two other layers, each formed from a different viral protein, VP7 and VP3 (30). The VP3 layer, together with the transcription complex of three minor enzymatic proteins, VP1, VP4, and VP6, and the 10 double-stranded RNA segments, forms the subcore, which is the first assembled particle that acts as the foundation for virus assembly. Expression of VP3 (103 kDa) using a heterologous baculovirus expression system results in spontaneous assembly of a particu...

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“…For instance, a maximum of 21 aa from HPV16 E7 was inserted into hepatitis B core antigen particles (31), and similar size constraints have been noted for HPV L1 (33). The insertion of a larger polypeptide, the 147-aa hen egg white lysozyme protein, into parvovirus capsids was presumably accomplished because it was fused to VP1, the parvovirus minor capsid protein (32).…”

Section: Discussionmentioning

confidence: 99%

“…Chimeric VLPs incorporating foreign antigens have been generated previously by using capsid proteins from other viruses, including parvovirus, HIV-1, and hepatitis virus (30)(31)(32), but they have not been shown to generate antitumor immunity. Furthermore, the foreign peptides have usually been inserted into the major capsid protein, whose ability to self-assemble limited the size of the insert to relatively small peptides.…”

Section: Discussionmentioning

confidence: 99%

Chimeric papillomavirus virus-like particles elicit antitumor immunity against the E7 oncoprotein in an HPV16 tumor model

Greenstone

Nieland

Visser

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

283

176

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Papillomavirus-like particles (VLPs) are a promising prophylactic vaccine candidate to prevent human papillomavirus (HPV) infections and associated epithelial neoplasia. However, they are unlikely to have therapeutic effects because the virion capsid proteins are not detected in the proliferating cells of the infected epithelia or in cervical carcinomas. To increase the number of viral antigen targets for cell-mediated immune responses in a VLP-based vaccine, we have generated stable chimeric VLPs consisting of the L1 major capsid protein plus the entire E7 (11 kDa) or E2 (43 kDa) nonstructural papillomavirus protein fused to the L2 minor capsid protein. The chimeric VLPs are indistinguishable from the parental VLPs in their morphology and in their ability to agglutinate erythrocytes and elicit high titers of neutralizing antibodies. Protection from tumor challenge was tested in C57BL͞6 mice by using the tumor cell line TC-1, which expresses HPV16 E7, but not the virion structural proteins. Injection of HPV16 L1͞L2-HPV16 E7 chimeric VLPs, but not HPV16 L1͞L2 VLPs, protected the mice from tumor challenge, even in the absence of adjuvant. The chimeric VLPs also induced protection against tumor challenge in major histocompatibility class IIdeficient mice, but not in ␤ 2 -microglobulin or perforin knockout mice implying that protection was mediated by class I-restricted cytotoxic lymphocytes. These findings raise the possibility that VLPs may generally be efficient vehicles for generating cellmediated immune responses and that, specifically, chimeric VLPs containing papillomavirus nonstructural proteins may increase the therapeutic potential of VLP-based prophylactic vaccines in humans.Human papillomaviruses (HPVs) that infect the genital tract are associated with human anogenital tract cancer, particularly cervical cancer (reviewed in ref. 1). HPVs are thought to be the primary causative agent in Ͼ90% of cervical cancers (2), with HPV16 being the type most frequently found in these tumors. Approximately 500,000 women develop cervical cancer each year, and 200,000 women die from it, making this disease the second-most common cause of cancer deaths in women worldwide (3).Significant advances have been made recently in the development of a candidate prophylactic vaccine against papillomavirus infections (reviewed in ref. 4). Expression of the papillomavirus major capsid protein, L1, in eukaryotic cells leads to self-assembly into virus-like particles (VLPs) that are morphologically indistinguishable from native virions and present the conformational epitopes required for the induction of high titer neutralizing antisera (5). L2, the minor capsid protein, coassembles with L1 at a ratio of Ϸ1 L2 molecule to 30 L1 molecules (6). Although L2 presents some epitopes that induce the production of neutralizing antiserum (7), most neutralizing antibodies induced by L1͞L2 VLPs recognize L1 determinants (8). Several studies have shown that L1 and L1͞L2 VLP-based vaccines protect animals against high dose experimental pap...

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Parvovirus particles as platforms for protein presentation.

Cited by 39 publications

References 34 publications

Conformational Changes in the VP1-Unique Region of Native Human Parvovirus B19 Lead to Exposure of Internal Sequences That Play a Role in Virus Neutralization and Infectivity

Conformational Changes in the VP1-Unique Region of Native Human Parvovirus B19 Lead to Exposure of Internal Sequences That Play a Role in Virus Neutralization and Infectivity

Assembly and Intracellular Localization of the Bluetongue Virus Core Protein VP3

Chimeric papillomavirus virus-like particles elicit antitumor immunity against the E7 oncoprotein in an HPV16 tumor model

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