Rotavirus structure: interactions between the structural proteins

Shaw, Andrea L.; Rothnagel, Rosalba; Zeng, Chi; Lawton, Jeffrey A.; Ramig, Robert F.; Estes, Mary K.; Prasad, B. V. Venkataram

doi:10.1007/978-3-7091-6553-9_3

Cited by 15 publications

(17 citation statements)

References 20 publications

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“…The expressed fusion protein contained a thrombin recognition site between VP8* and GST, for removal of the GST portion. and corresponded to the expected VP8* cleavage fragment (beginning at the thrombin recognition site, with N-terminal sequence GSPGIPD), as well as nonspecific cleavage products, one with N-terminal sequence Y 16 SVDLHDE, and smaller unidentifiable fragments. The nonspecific cleavage of the fusion protein may also be responsible for the lower than expected VP8* yield, as any of the VP8* cleaved prematurely from GST would have not bound to the GS4B, and therefore washed from the column.…”

Section: Figmentioning

confidence: 88%

“…Of these 11 gene segments, six code for structural proteins, and five code for nonstructural proteins. The six structural proteins are arranged into three concentric layers surrounding the rotavirus genome (16). The outermost layer is composed of two proteins, VP7 and VP4.…”

mentioning

confidence: 99%

“…The minor component of the outer shell, VP4, is present as a series of spikes that project outward from the VP7 shell. VP4 is nonglycosylated, has a molecular weight of 88 kDa, and constitutes 1.5% of the virus protein (1,16). VP4 has been implicated in several important functions, including cell attachment and penetration, hemagglutination, neutralization, host range, and virulence (1,17), and viral infectivity is enhanced by the cleavage of VP4, by trypsin, into VP5 (the 60-kDa C-terminal segment) and VP8 (the 28-kDa N-terminal segment) (18).…”

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confidence: 99%

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Cloning and Expression of Human Rotavirus Spike Protein, VP8*, in Escherichia coli

Kovacs-Nolan

Sasaki

Yoo

et al. 2001

Biochemical and Biophysical Research Communications

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Section: Figmentioning

confidence: 88%

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confidence: 99%

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confidence: 99%

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Cloning and Expression of Human Rotavirus Spike Protein, VP8*, in Escherichia coli

Kovacs-Nolan

Sasaki

Yoo

et al. 2001

Biochemical and Biophysical Research Communications

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“…Structural and biochemical analyses show that rotaviruses are large, icosahedral particles consisting of three concentric capsid layers surrounding a genome of 11 segments of double-stranded RNA (15,45). VP4 is a nonglycosylated protein and forms spikes that project from the rotavirus surface.…”

mentioning

confidence: 99%

Interactions of Rotavirus VP4 Spike Protein with the Endosomal Protein Rab5 and the Prenylated Rab Acceptor PRA1

Enouf

Chwetzoff

Trugnan

et al. 2003

J Virol

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Rotavirus spike protein VP4 is implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, virulence, and host range. It is present at the plasma membrane and colocalizes with the cytoskeleton in infected cells. We looked for cellular partners responsible for the localization of VP4 by two-hybrid screening of a monkey CV1 cell cDNA library. In the screen we isolated repeatedly three cDNAs encoding either two isoforms (a and c) of Rab5 protein or the prenylated Rab acceptor (PRA1). The small GTPase Rab5 is a molecule regulating the vesicular traffic and the motility of early endosomes along microtubules. Rab5 interacts with a large number of effectors, in particular with PRA1. Interactions of VP4 with both partners, Rab5 and PRA1, were confirmed by coimmunoprecipitation from infected-or transfectedcell lysates. Interaction of Rab5 and PRA1 was restricted to free VP4, since neither triple-layered particles nor NSP4-VP4-VP7 heterotrimeric complexes could be coprecipitated. Site-directed and deletion mutants of VP4 were used to map a VP4 domain(s) interacting with Rab5 or PRA1. Of the 10 mutants tested, 2 interacted exclusively with a single partner. In contrast, the domain extending from amino acids 560 to 722 of VP4 is essential for both interactions. These results suggest that Rab5 and PRA1 may be involved in the localization and trafficking of VP4 in infected cells.Rotaviruses are the leading cause of severe gastroenteritis in young children worldwide (24). Structural and biochemical analyses show that rotaviruses are large, icosahedral particles consisting of three concentric capsid layers surrounding a genome of 11 segments of double-stranded RNA (15, 45). VP4 is a nonglycosylated protein and forms spikes that project from the rotavirus surface. Trypsin cleavage of VP4 into the fragments VP8* and VP5* is required for viral infectivity and stabilization of the spikes (11, 30). VP4 has been implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, and virulence (10,17,26,32). It has been shown previously that VP5*, which includes a conserved hydrophobic region located between amino acids (aa) 384 and 401 of VP4, is a specific membrane-permeabilizing protein and could play a role in the cellular entry of rotaviruses (12). Heterotrimers consisting of VP4, NSP4, and VP7 may participate in the budding of the single-shelled particles into the lumen of the endoplasmic reticulum, where maturation to double-shelled particles seems to occur (33, 39).Rab proteins are a family of Ras-related low-molecularweight monomeric GTP-binding proteins (ϳ20 to 29 kDa) which are key regulators of vesicular transport within eukaryotic cells (7). Several effector proteins alter either the state of phosphorylation or the intracytoplasmic location of Rab proteins and thus modulate their biological activity (25). The first modification refers to a posttranslational covalent addition of two 20-carbon isoprenoid geranyl-geranyl gro...

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“…They are responsible for more than 850,000 deaths per year (14). As a member of the Reoviridae family, rotavirus has a segmented double-stranded RNA genome, enclosed in a viral capsid constituted of three concentric protein layers (37). Electron microscopy studies show that viral morphogenesis begins in cytoplasmic inclusions, termed viroplasms, where the central core and single-shelled particles are assembled (3,10).…”

mentioning

confidence: 99%

Rotavirus Spike Protein VP4 Is Present at the Plasma Membrane and Is Associated with Microtubules in Infected Cells

Nejmeddine

Trugnan

Sapin

et al. 2000

J Virol

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VP4 is an unglycosylated protein of the outer layer of the capsid of rotavirus. It forms spikes that project from the outer layer of mature virions, which is mainly constituted by glycoprotein VP7. VP4 has been implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, virulence, and host range. Previous studies indicated that VP4 is located in the space between the periphery of the viroplasm and the outside of the endoplasmic reticulum in rotavirus-infected cells. Confocal microscopy of infected MA104 monolayers, immunostained with specific monoclonal antibodies, revealed that a significant fraction of VP4 was present at the plasma membrane early after infection. Another fraction of VP4 is cytoplasmic and colocalizes with ␤-tubulin. Flow cytometry analysis confirmed that at the early stage of viral infection, VP4 was present on the plasma membrane and that its N-terminal region, the VP8* subunit, was accessible to antibodies. Biotin labeling of the infected cell surface monolayer with a cell-impermeable reagent allowed the identification of the noncleaved form of VP4 that was associated with the glycoprotein VP7. The localization of VP4 was not modified in cells transfected with a plasmid allowing the expression of a fusion protein consisting of VP4 and the green fluorescent protein. The present data suggest that VP4 reaches the plasma membrane through the microtubule network and that other viral proteins are dispensable for its targeting and transport.Rotaviruses are the most important etiologic agents of severe dehydrating infantile gastroenteritis in developed and developing countries (17). They are responsible for more than 850,000 deaths per year (14). As a member of the Reoviridae family, rotavirus has a segmented double-stranded RNA genome, enclosed in a viral capsid constituted of three concentric protein layers (37). Electron microscopy studies show that viral morphogenesis begins in cytoplasmic inclusions, termed viroplasms, where the central core and single-shelled particles are assembled (3, 10). VP4 is an unglycosylated protein and forms spikes that project from the outer layer of mature virions, which is mainly constituted by the glycoprotein VP7 (1, 34). VP4 has been implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, virulence, and host range (5,12,18,23). It has been shown that the infectivity of rotaviruses is increased and is probably dependent on trypsin treatment of the virus (11). This proteolytic treatment results in the specific cleavage of VP4 to polypeptides VP8* and VP5*, which represent, respectively, the amino-and carboxyl-terminal regions of the protein (22). VP4 possesses a conserved hydrophobic region located between amino acids 384 and 401 that shares some homology with the internal fusion sites of Semliki Forest virus and Sindbis virus E1 spike proteins (25). Recently, it has been shown that VP5*, which includes this hydrophobic domain, is a specific membrane-p...

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Rotavirus structure: interactions between the structural proteins

Cited by 15 publications

References 20 publications

Cloning and Expression of Human Rotavirus Spike Protein, VP8*, in Escherichia coli

Cloning and Expression of Human Rotavirus Spike Protein, VP8*, in Escherichia coli

Interactions of Rotavirus VP4 Spike Protein with the Endosomal Protein Rab5 and the Prenylated Rab Acceptor PRA1

Rotavirus Spike Protein VP4 Is Present at the Plasma Membrane and Is Associated with Microtubules in Infected Cells

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