Switch-on of transcriptase function in reovirus: Analysis of polypeptide changes using 2-D gels

Ewing, Donald D.; Sargent, M. D.; Borsa, J.

doi:10.1016/0042-6822(85)90285-5

Cited by 14 publications

(12 citation statements)

References 17 publications

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“…These observations strongly suggest that the outer capsid in general, and protein 1 in particular, play a role in regulating the activity of the core-associated transcriptases. Other experiments showed that incubation of ISVPs with K ϩ or Cs ϩ ions at 37°C is sufficient for derepression of transcriptase activity and that proteolytic removal of 1 from viral particles is not required (9,12,23,57). In the present study, we found that incubation of ISVPs with Cs ϩ ions accelerates, in concert, both conformational change in 1 (Fig.…”

Section: Vol 76 2002 Entry-related Structural Transition In Reovirusupporting

confidence: 63%

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Strategy for Nonenveloped Virus Entry: a Hydrophobic Conformer of the Reovirus Membrane Penetration Protein μ1 Mediates Membrane Disruption

Chandran

Farsetta

Nibert

2002

J Virol

167

335

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The mechanisms employed by nonenveloped animal viruses to penetrate the membranes of their host cells remain enigmatic. Membrane penetration by the nonenveloped mammalian reoviruses is believed to deliver a partially uncoated, but still large (ϳ70-nm), particle with active transcriptases for viral mRNA synthesis directly into the cytoplasm. This process is likely initiated by a particle form that resembles infectious subvirion particles (ISVPs), disassembly intermediates produced from virions by proteolytic uncoating. Consistent with that idea, ISVPs, but not virions, can induce disruption of membranes in vitro. Both activities ascribed to ISVP-like particles, membrane disruption in vitro and membrane penetration within cells, are linked to N-myristoylated outer-capsid protein 1, present in 600 copies at the surfaces of ISVPs. To understand how 1 fulfills its role as the reovirus penetration protein, we monitored changes in ISVPs during the permeabilization of red blood cells induced by these particles. Hemolysis was preceded by a major structural transition in ISVPs, characterized by conformational change in 1 and elution of fibrous attachment protein 1. The altered conformer of 1 was required for hemolysis and was markedly hydrophobic. The structural transition in ISVPs was further accompanied by derepression of genome-dependent mRNA synthesis by the particle-associated transcriptases. We propose a model for reovirus entry in which (i) primed and triggered conformational changes, analogous to those in enveloped-virus fusion proteins, generate a hydrophobic 1 conformer capable of inserting into and disrupting cell membranes and (ii) activation of the viral particles for membrane interaction and mRNA synthesis are concurrent events. Reoviruses provide an opportune system for defining the molecular details of membrane penetration by a large nonenveloped animal virus.

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Section: Vol 76 2002 Entry-related Structural Transition In Reovirusupporting

confidence: 63%

“…8). A role for the 1-to-1* change in this process is certain, given previous evidence implicating M2 and 1 as determinants of both transcriptase activation and repression (21,23,24). How does 1* allow derepression of the viral transcription machines located within the inner capsid?…”

Section: Vol 76 2002 Entry-related Structural Transition In Reovirumentioning

confidence: 99%

Strategy for Nonenveloped Virus Entry: a Hydrophobic Conformer of the Reovirus Membrane Penetration Protein μ1 Mediates Membrane Disruption

Chandran

Farsetta

Nibert

2002

J Virol

167

335

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“…An infrequently cited paper by Ewing et al 46 reported loss of a small amount of "d 1 " protein in two-dimensional gels after transcriptase activation of ISVPs in the absence of protease. According to our interpretation and terminology, the d 1 protein is the same as the m1d fragment, and thus these previous authors appear to have been describing the same phenomenon as we are now showing in Figure 8.…”

Section: Discussionmentioning

confidence: 97%

Putative Autocleavage of Reovirus μ1 Protein in Concert with Outer-capsid Disassembly and Activation for Membrane Permeabilization

Nibert

Odegard

Agosto

et al. 2005

Journal of Molecular Biology

116

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“…A strain difference in the treatment conditions needed to generate cores and activate transcript elongation was mapped to the M2 gene, suggesting that 1 plays a regulatory role in the switch-on of transcriptase activity (15). According to a current model, virions undergo transcriptase activation via two steps (7,21,43). In the first FIG.…”

Section: Discussionmentioning

confidence: 99%

Protease Cleavage of Reovirus Capsid Protein μ1/μ1C Is Blocked by Alkyl Sulfate Detergents, Yielding a New Type of Infectious Subvirion Particle

Chandran

Nibert

1998

J Virol

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Mammalian reovirus virions undergo partial disassembly of the outer capsid upon exposure to proteases in vitro, producing infectious subvirion particles (ISVPs) that lack protein ς3 and contain protein μ1/μ1C as endoprotease-generated fragments μ1δ/δ and φ. ISVPs are thought to be required for two early steps in reovirus infection: membrane penetration and activation of the particle-bound viral transcriptase complexes. Genetic and biochemical evidence implicates outer-capsid protein μ1 in both these steps. To determine whether the cleavage of μ1/μ1C is relevant to the unique properties of ISVPs, we analyzed the properties of novel subvirion particles that lacked ς3 yet retained μ1/μ1C in an uncleaved but cleavable form. These detergent-plus-protease subvirion particles (dpSVPs) were produced by treating virions with chymotrypsin in the presence of micelle-forming concentrations of alkyl sulfate detergents. Infections with dpSVPs in murine L or canine MDCK cells provided evidence that the cleavage of μ1/μ1C during viral entry into these cells is dispensable for reovirus infection. Additionally, dpSVPs behaved like ISVPs in their capacity to permeabilize lipid bilayers and to undergo transcriptase activation in vitro, supporting the conclusion that cleavage of μ1/μ1C to μ1δ/δ and φ during viral entry is not required for either membrane penetration or transcriptase activation in cells. The capacity of alkyl sulfate detergents to inhibit the cleavage of μ1/μ1C in a reversible fashion suggests a specific association between virus particle and detergent micelles that may mimic virus particle-phospholipid membrane interactions during reovirus entry into cells.

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Switch-on of transcriptase function in reovirus: Analysis of polypeptide changes using 2-D gels

Cited by 14 publications

References 17 publications

Strategy for Nonenveloped Virus Entry: a Hydrophobic Conformer of the Reovirus Membrane Penetration Protein μ1 Mediates Membrane Disruption

Strategy for Nonenveloped Virus Entry: a Hydrophobic Conformer of the Reovirus Membrane Penetration Protein μ1 Mediates Membrane Disruption

Putative Autocleavage of Reovirus μ1 Protein in Concert with Outer-capsid Disassembly and Activation for Membrane Permeabilization

Protease Cleavage of Reovirus Capsid Protein μ1/μ1C Is Blocked by Alkyl Sulfate Detergents, Yielding a New Type of Infectious Subvirion Particle

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