The Roles of Template RNA and Replication Proteins in the Formation of Semliki Forest Virus Replication Spherules

Research output: ThesisDoctoral ThesisCollection of Articles

Abstract

All positive-strand RNA viruses replicate their RNA genomes in close association with cellular membranes. A great variety of cellular membranes are utilized by different viruses and those membranes are extensively modified to support viral replication and to protect the viral RNA from host cell defense mechanisms. Alphaviruses, including Semliki Forest virus (SFV), are positive-strand RNA viruses replicating their RNA on membranes derived from endosomal and lysosomal compartments. SFV induces small invaginations called spherules on plasma membrane and on endosomal membranes. Viral replication complex assembly, spherule formation and initiation of replication are carefully orchestrated events and are guided by specific sequence elements within the genomic RNA as well as by important enzymatic activities of nonstructural proteins (nsPs). The aim of this research was to study in detail how alphavirus replication complexes are assembled and to define the minimum requirements for spherule formation by using a plasmid-derived transreplication system mimicking SFV replication. The role of the genomic RNA in replication was deciphered by using RNA templates, which were either modified or differed in length. Use of RNA templates differing in length clearly showed that they define the spherule diameter suggesting that the template has a significant role in spherule formation. By modifying or deleting specific sequences from the template it was shown that highly conserved RNA elements are important for SFV replication and do not tolerate modifications without compromising replication. Study with the nsPs of SFV showed that the enzymatic activities essential for virus replication are also needed for spherule formation and that enzymes like helicase, protease and polymerase are absolutely essential for replication. Membrane association of the replication complex is also required to establish virus replication in the cells. The work with mutated nonstructural proteins and modified templates revealed a clear correlation between the minus-strand synthesis and spherule formation. This work describes the alphavirus replication processes in detail and provides new principles, which may be generally applicable to study the positive-sense RNA virus replication and the formation of virus-induced membranous replication spherules.
Original languageEnglish
Place of PublicationHelsinki
Publisher
Print ISBNs978-951-51-2081-6
Electronic ISBNs978-951-51-2082-3
Publication statusPublished - 20 May 2016
MoE publication typeG5 Doctoral dissertation (article)

Fields of Science

  • 1183 Plant biology, microbiology, virology

Cite this

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title = "The Roles of Template RNA and Replication Proteins in the Formation of Semliki Forest Virus Replication Spherules",
abstract = "All positive-strand RNA viruses replicate their RNA genomes in close association with cellular membranes. A great variety of cellular membranes are utilized by different viruses and those membranes are extensively modified to support viral replication and to protect the viral RNA from host cell defense mechanisms. Alphaviruses, including Semliki Forest virus (SFV), are positive-strand RNA viruses replicating their RNA on membranes derived from endosomal and lysosomal compartments. SFV induces small invaginations called spherules on plasma membrane and on endosomal membranes. Viral replication complex assembly, spherule formation and initiation of replication are carefully orchestrated events and are guided by specific sequence elements within the genomic RNA as well as by important enzymatic activities of nonstructural proteins (nsPs). The aim of this research was to study in detail how alphavirus replication complexes are assembled and to define the minimum requirements for spherule formation by using a plasmid-derived transreplication system mimicking SFV replication. The role of the genomic RNA in replication was deciphered by using RNA templates, which were either modified or differed in length. Use of RNA templates differing in length clearly showed that they define the spherule diameter suggesting that the template has a significant role in spherule formation. By modifying or deleting specific sequences from the template it was shown that highly conserved RNA elements are important for SFV replication and do not tolerate modifications without compromising replication. Study with the nsPs of SFV showed that the enzymatic activities essential for virus replication are also needed for spherule formation and that enzymes like helicase, protease and polymerase are absolutely essential for replication. Membrane association of the replication complex is also required to establish virus replication in the cells. The work with mutated nonstructural proteins and modified templates revealed a clear correlation between the minus-strand synthesis and spherule formation. This work describes the alphavirus replication processes in detail and provides new principles, which may be generally applicable to study the positive-sense RNA virus replication and the formation of virus-induced membranous replication spherules.",
keywords = "1183 Plant biology, microbiology, virology",
author = "Kallio, {Katri Anneli}",
year = "2016",
month = "5",
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language = "English",
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series = "Dissertationes scholae doctoralis ad sanitatem investigandam Universitatis Helsinkiensis",
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The Roles of Template RNA and Replication Proteins in the Formation of Semliki Forest Virus Replication Spherules. / Kallio, Katri Anneli.

Helsinki : Helsingin yliopisto, 2016. 54 p.

Research output: ThesisDoctoral ThesisCollection of Articles

TY - THES

T1 - The Roles of Template RNA and Replication Proteins in the Formation of Semliki Forest Virus Replication Spherules

AU - Kallio, Katri Anneli

PY - 2016/5/20

Y1 - 2016/5/20

N2 - All positive-strand RNA viruses replicate their RNA genomes in close association with cellular membranes. A great variety of cellular membranes are utilized by different viruses and those membranes are extensively modified to support viral replication and to protect the viral RNA from host cell defense mechanisms. Alphaviruses, including Semliki Forest virus (SFV), are positive-strand RNA viruses replicating their RNA on membranes derived from endosomal and lysosomal compartments. SFV induces small invaginations called spherules on plasma membrane and on endosomal membranes. Viral replication complex assembly, spherule formation and initiation of replication are carefully orchestrated events and are guided by specific sequence elements within the genomic RNA as well as by important enzymatic activities of nonstructural proteins (nsPs). The aim of this research was to study in detail how alphavirus replication complexes are assembled and to define the minimum requirements for spherule formation by using a plasmid-derived transreplication system mimicking SFV replication. The role of the genomic RNA in replication was deciphered by using RNA templates, which were either modified or differed in length. Use of RNA templates differing in length clearly showed that they define the spherule diameter suggesting that the template has a significant role in spherule formation. By modifying or deleting specific sequences from the template it was shown that highly conserved RNA elements are important for SFV replication and do not tolerate modifications without compromising replication. Study with the nsPs of SFV showed that the enzymatic activities essential for virus replication are also needed for spherule formation and that enzymes like helicase, protease and polymerase are absolutely essential for replication. Membrane association of the replication complex is also required to establish virus replication in the cells. The work with mutated nonstructural proteins and modified templates revealed a clear correlation between the minus-strand synthesis and spherule formation. This work describes the alphavirus replication processes in detail and provides new principles, which may be generally applicable to study the positive-sense RNA virus replication and the formation of virus-induced membranous replication spherules.

AB - All positive-strand RNA viruses replicate their RNA genomes in close association with cellular membranes. A great variety of cellular membranes are utilized by different viruses and those membranes are extensively modified to support viral replication and to protect the viral RNA from host cell defense mechanisms. Alphaviruses, including Semliki Forest virus (SFV), are positive-strand RNA viruses replicating their RNA on membranes derived from endosomal and lysosomal compartments. SFV induces small invaginations called spherules on plasma membrane and on endosomal membranes. Viral replication complex assembly, spherule formation and initiation of replication are carefully orchestrated events and are guided by specific sequence elements within the genomic RNA as well as by important enzymatic activities of nonstructural proteins (nsPs). The aim of this research was to study in detail how alphavirus replication complexes are assembled and to define the minimum requirements for spherule formation by using a plasmid-derived transreplication system mimicking SFV replication. The role of the genomic RNA in replication was deciphered by using RNA templates, which were either modified or differed in length. Use of RNA templates differing in length clearly showed that they define the spherule diameter suggesting that the template has a significant role in spherule formation. By modifying or deleting specific sequences from the template it was shown that highly conserved RNA elements are important for SFV replication and do not tolerate modifications without compromising replication. Study with the nsPs of SFV showed that the enzymatic activities essential for virus replication are also needed for spherule formation and that enzymes like helicase, protease and polymerase are absolutely essential for replication. Membrane association of the replication complex is also required to establish virus replication in the cells. The work with mutated nonstructural proteins and modified templates revealed a clear correlation between the minus-strand synthesis and spherule formation. This work describes the alphavirus replication processes in detail and provides new principles, which may be generally applicable to study the positive-sense RNA virus replication and the formation of virus-induced membranous replication spherules.

KW - 1183 Plant biology, microbiology, virology

M3 - Doctoral Thesis

SN - 978-951-51-2081-6

T3 - Dissertationes scholae doctoralis ad sanitatem investigandam Universitatis Helsinkiensis

PB - Helsingin yliopisto

CY - Helsinki

ER -

Kallio KA. The Roles of Template RNA and Replication Proteins in the Formation of Semliki Forest Virus Replication Spherules. Helsinki: Helsingin yliopisto, 2016. 54 p. (Dissertationes scholae doctoralis ad sanitatem investigandam Universitatis Helsinkiensis; 25).