The nsp2 replicase proteins of murine hepatitis virus and severe acute respiratory syndrome coronavirus are dispensable for viral replication.
Identifieur interne : 001208 ( Ncbi/Checkpoint ); précédent : 001207; suivant : 001209The nsp2 replicase proteins of murine hepatitis virus and severe acute respiratory syndrome coronavirus are dispensable for viral replication.
Auteurs : Rachel L. Graham [États-Unis] ; Amy C. Sims ; Sarah M. Brockway ; Ralph S. Baric ; Mark R. DenisonSource :
- Journal of virology [ 0022-538X ] ; 2005.
Descripteurs français
- KwdFr :
- ARN viral (biosynthèse), Animaux, Cadres ouverts de lecture (génétique), Délétion de gène, Lignée cellulaire, Polyprotéines (génétique), Protéines virales non structurales (physiologie), RNA replicase (physiologie), Réplication virale, Test de complémentation, Virus de l'hépatite murine (génétique), Virus de l'hépatite murine (physiologie), Virus du SRAS (génétique), Virus du SRAS (physiologie).
- MESH :
- biosynthèse : ARN viral.
- génétique : Cadres ouverts de lecture, Polyprotéines, Virus de l'hépatite murine, Virus du SRAS.
- physiologie : Protéines virales non structurales, RNA replicase, Virus de l'hépatite murine, Virus du SRAS.
- Animaux, Délétion de gène, Lignée cellulaire, Réplication virale, Test de complémentation.
English descriptors
- KwdEn :
- Animals, Cell Line, Gene Deletion, Genetic Complementation Test, Murine hepatitis virus (genetics), Murine hepatitis virus (physiology), Open Reading Frames (genetics), Polyproteins (genetics), RNA Replicase (physiology), RNA, Viral (biosynthesis), SARS Virus (genetics), SARS Virus (physiology), Viral Nonstructural Proteins (physiology), Virus Replication.
- MESH :
- chemical , biosynthesis : RNA, Viral.
- chemical , genetics : Polyproteins.
- genetics : Murine hepatitis virus, Open Reading Frames, SARS Virus.
- physiology : Murine hepatitis virus, RNA Replicase, SARS Virus, Viral Nonstructural Proteins.
- Animals, Cell Line, Gene Deletion, Genetic Complementation Test, Virus Replication.
Abstract
The positive-stranded RNA genome of the coronaviruses is translated from ORF1 to yield polyproteins that are proteolytically processed into intermediate and mature nonstructural proteins (nsps). Murine hepatitis virus (MHV) and severe acute respiratory syndrome coronavirus (SARS-CoV) polyproteins incorporate 16 protein domains (nsps), with nsp1 and nsp2 being the most variable among the coronaviruses and having no experimentally confirmed or predicted functions in replication. To determine if nsp2 is essential for viral replication, MHV and SARS-CoV genome RNA was generated with deletions of the nsp2 coding sequence (MHVDeltansp2 and SARSDeltansp2, respectively). Infectious MHVDeltansp2 and SARSDeltansp2 viruses recovered from electroporated cells had 0.5 to 1 log10 reductions in peak titers in single-cycle growth assays, as well as a reduction in viral RNA synthesis that was not specific for any positive-stranded RNA species. The Deltansp2 mutant viruses lacked expression of both nsp2 and an nsp2-nsp3 precursor, but cleaved the engineered chimeric nsp1-nsp3 cleavage site as efficiently as the native nsp1-nsp2 cleavage site. Replication complexes in MHVDeltansp2-infected cells lacked nsp2 but were morphologically indistinguishable from those of wild-type MHV by immunofluorescence. nsp2 expressed in cells by stable retroviral transduction was specifically recruited to viral replication complexes upon infection with MHVDeltansp2. These results demonstrate that while nsp2 of MHV and SARS-CoV is dispensable for viral replication in cell culture, deletion of the nsp2 coding sequence attenuates viral growth and RNA synthesis. These findings also provide a system for the study of determinants of nsp targeting and function.
DOI: 10.1128/JVI.79.21.13399-13411.2005
PubMed: 16227261
Affiliations:
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Graham</name><affiliation wicri:level="2"><nlm:affiliation>Department of Pediatrics, Vanderbilt University Medical Center, D6217 MCN, Nashville, TN 37232-2581, USA.</nlm:affiliation><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Pediatrics, Vanderbilt University Medical Center, D6217 MCN, Nashville, TN 37232-2581</wicri:regionArea><placeName><region type="state">Tennessee</region></placeName></affiliation></author><author><name sortKey="Sims, Amy C" sort="Sims, Amy C" uniqKey="Sims A" first="Amy C" last="Sims">Amy C. Sims</name></author><author><name sortKey="Brockway, Sarah M" sort="Brockway, Sarah M" uniqKey="Brockway S" first="Sarah M" last="Brockway">Sarah M. Brockway</name></author><author><name sortKey="Baric, Ralph S" sort="Baric, Ralph S" uniqKey="Baric R" first="Ralph S" last="Baric">Ralph S. Baric</name></author><author><name sortKey="Denison, Mark R" sort="Denison, Mark R" uniqKey="Denison M" first="Mark R" last="Denison">Mark R. Denison</name></author></analytic><series><title level="j">Journal of virology</title><idno type="ISSN">0022-538X</idno><imprint><date when="2005" type="published">2005</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>Cell Line</term><term>Gene Deletion</term><term>Genetic Complementation Test</term><term>Murine hepatitis virus (genetics)</term><term>Murine hepatitis virus (physiology)</term><term>Open Reading Frames (genetics)</term><term>Polyproteins (genetics)</term><term>RNA Replicase (physiology)</term><term>RNA, Viral (biosynthesis)</term><term>SARS Virus (genetics)</term><term>SARS Virus (physiology)</term><term>Viral Nonstructural Proteins (physiology)</term><term>Virus Replication</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral (biosynthèse)</term><term>Animaux</term><term>Cadres ouverts de lecture (génétique)</term><term>Délétion de gène</term><term>Lignée cellulaire</term><term>Polyprotéines (génétique)</term><term>Protéines virales non structurales (physiologie)</term><term>RNA replicase (physiologie)</term><term>Réplication virale</term><term>Test de complémentation</term><term>Virus de l'hépatite murine (génétique)</term><term>Virus de l'hépatite murine (physiologie)</term><term>Virus du SRAS (génétique)</term><term>Virus du SRAS (physiologie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>RNA, Viral</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Polyproteins</term></keywords><keywords scheme="MESH" qualifier="biosynthèse" xml:lang="fr"><term>ARN viral</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Murine hepatitis virus</term><term>Open Reading Frames</term><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Cadres ouverts de lecture</term><term>Polyprotéines</term><term>Virus de l'hépatite murine</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="physiologie" xml:lang="fr"><term>Protéines virales non structurales</term><term>RNA replicase</term><term>Virus de l'hépatite murine</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Murine hepatitis virus</term><term>RNA Replicase</term><term>SARS Virus</term><term>Viral Nonstructural Proteins</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Cell Line</term><term>Gene Deletion</term><term>Genetic Complementation Test</term><term>Virus Replication</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Animaux</term><term>Délétion de gène</term><term>Lignée cellulaire</term><term>Réplication virale</term><term>Test de complémentation</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The positive-stranded RNA genome of the coronaviruses is translated from ORF1 to yield polyproteins that are proteolytically processed into intermediate and mature nonstructural proteins (nsps). Murine hepatitis virus (MHV) and severe acute respiratory syndrome coronavirus (SARS-CoV) polyproteins incorporate 16 protein domains (nsps), with nsp1 and nsp2 being the most variable among the coronaviruses and having no experimentally confirmed or predicted functions in replication. To determine if nsp2 is essential for viral replication, MHV and SARS-CoV genome RNA was generated with deletions of the nsp2 coding sequence (MHVDeltansp2 and SARSDeltansp2, respectively). Infectious MHVDeltansp2 and SARSDeltansp2 viruses recovered from electroporated cells had 0.5 to 1 log10 reductions in peak titers in single-cycle growth assays, as well as a reduction in viral RNA synthesis that was not specific for any positive-stranded RNA species. The Deltansp2 mutant viruses lacked expression of both nsp2 and an nsp2-nsp3 precursor, but cleaved the engineered chimeric nsp1-nsp3 cleavage site as efficiently as the native nsp1-nsp2 cleavage site. Replication complexes in MHVDeltansp2-infected cells lacked nsp2 but were morphologically indistinguishable from those of wild-type MHV by immunofluorescence. nsp2 expressed in cells by stable retroviral transduction was specifically recruited to viral replication complexes upon infection with MHVDeltansp2. These results demonstrate that while nsp2 of MHV and SARS-CoV is dispensable for viral replication in cell culture, deletion of the nsp2 coding sequence attenuates viral growth and RNA synthesis. These findings also provide a system for the study of determinants of nsp targeting and function.</div></front></TEI><affiliations><list><country><li>États-Unis</li></country><region><li>Tennessee</li></region></list><tree><noCountry><name sortKey="Baric, Ralph S" sort="Baric, Ralph S" uniqKey="Baric R" first="Ralph S" last="Baric">Ralph S. Baric</name><name sortKey="Brockway, Sarah M" sort="Brockway, Sarah M" uniqKey="Brockway S" first="Sarah M" last="Brockway">Sarah M. Brockway</name><name sortKey="Denison, Mark R" sort="Denison, Mark R" uniqKey="Denison M" first="Mark R" last="Denison">Mark R. Denison</name><name sortKey="Sims, Amy C" sort="Sims, Amy C" uniqKey="Sims A" first="Amy C" last="Sims">Amy C. Sims</name></noCountry><country name="États-Unis"><region name="Tennessee"><name sortKey="Graham, Rachel L" sort="Graham, Rachel L" uniqKey="Graham R" first="Rachel L" last="Graham">Rachel L. Graham</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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