The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein.
Identifieur interne : 003573 ( Main/Exploration ); précédent : 003572; suivant : 003574The nonstructural protein 8 (nsp8) of the SARS coronavirus interacts with its ORF6 accessory protein.
Auteurs : Purnima Kumar [Inde] ; Vithiagaran Gunalan ; Boping Liu ; Vincent T K. Chow ; Julian Druce ; Chris Birch ; Mike Catton ; Burtram C. Fielding ; Yee-Joo Tan ; Sunil K. LalSource :
- Virology [ 0042-6822 ] ; 2007.
Descripteurs français
- KwdFr :
- Animaux, Clonage moléculaire, Cytoplasme (), Immunoprécipitation, Liaison aux protéines, Lignée cellulaire, Microscopie confocale, Microscopie de fluorescence, Protéines virales non structurales (génétique), Protéines virales non structurales (métabolisme), Techniques de double hybride, Virus du SRAS (métabolisme).
- MESH :
English descriptors
- KwdEn :
- Animals, Cell Line, Chlorocebus aethiops, Cloning, Molecular, Cytoplasm (chemistry), Immunoprecipitation, Microscopy, Confocal, Microscopy, Fluorescence, Protein Binding, SARS Virus (metabolism), Two-Hybrid System Techniques, Viral Nonstructural Proteins (genetics), Viral Nonstructural Proteins (metabolism).
- MESH :
- chemical , genetics : Viral Nonstructural Proteins.
- chemistry : Cytoplasm.
- metabolism : SARS Virus, Viral Nonstructural Proteins.
- Animals, Cell Line, Chlorocebus aethiops, Cloning, Molecular, Immunoprecipitation, Microscopy, Confocal, Microscopy, Fluorescence, Protein Binding, Two-Hybrid System Techniques.
Abstract
Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) caused a severe outbreak in several regions of the world in 2003. The SARS-CoV genome is predicted to contain 14 functional open reading frames (ORFs). The first ORF (1a and 1b) encodes a large polyprotein that is cleaved into nonstructural proteins (nsp). The other ORFs encode for four structural proteins (spike, membrane, nucleocapsid and envelope) as well as eight SARS-CoV-specific accessory proteins (3a, 3b, 6, 7a, 7b, 8a, 8b and 9b). In this report we have cloned the predicted nsp8 gene and the ORF6 gene of the SARS-CoV and studied their abilities to interact with each other. We expressed the two proteins as fusion proteins in the yeast two-hybrid system to demonstrate protein-protein interactions and tested the same using a yeast genetic cross. Further the strength of the interaction was measured by challenging growth of the positive interaction clones on increasing gradients of 2-amino trizole. The interaction was then verified by expressing both proteins separately in-vitro in a coupled-transcription translation system and by coimmunoprecipitation in mammalian cells. Finally, colocalization experiments were performed in SARS-CoV infected Vero E6 mammalian cells to confirm the nsp8-ORF6 interaction. To the best of our knowledge, this is the first report of the interaction between a SARS-CoV accessory protein and nsp8 and our findings suggest that ORF6 protein may play a role in virus replication.
DOI: 10.1016/j.virol.2007.04.029
PubMed: 17532020
Affiliations:
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O. Box: 10504, Aruna Asaf Ali Road, New Delhi 110067, India.</nlm:affiliation><country xml:lang="fr">Inde</country><wicri:regionArea>Virology Group, International Centre for Genetic Engineering and Biotechnology, P. O. Box: 10504, Aruna Asaf Ali Road, New Delhi 110067</wicri:regionArea><wicri:noRegion>New Delhi 110067</wicri:noRegion></affiliation></author><author><name sortKey="Gunalan, Vithiagaran" sort="Gunalan, Vithiagaran" uniqKey="Gunalan V" first="Vithiagaran" last="Gunalan">Vithiagaran Gunalan</name></author><author><name sortKey="Liu, Boping" sort="Liu, Boping" uniqKey="Liu B" first="Boping" last="Liu">Boping Liu</name></author><author><name sortKey="Chow, Vincent T K" sort="Chow, Vincent T K" uniqKey="Chow V" first="Vincent T K" last="Chow">Vincent T K. Chow</name></author><author><name sortKey="Druce, Julian" sort="Druce, Julian" uniqKey="Druce J" first="Julian" last="Druce">Julian Druce</name></author><author><name sortKey="Birch, Chris" sort="Birch, Chris" uniqKey="Birch C" first="Chris" last="Birch">Chris Birch</name></author><author><name sortKey="Catton, Mike" sort="Catton, Mike" uniqKey="Catton M" first="Mike" last="Catton">Mike Catton</name></author><author><name sortKey="Fielding, Burtram C" sort="Fielding, Burtram C" uniqKey="Fielding B" first="Burtram C" last="Fielding">Burtram C. Fielding</name></author><author><name sortKey="Tan, Yee Joo" sort="Tan, Yee Joo" uniqKey="Tan Y" first="Yee-Joo" last="Tan">Yee-Joo Tan</name></author><author><name sortKey="Lal, Sunil K" sort="Lal, Sunil K" uniqKey="Lal S" first="Sunil K" last="Lal">Sunil K. Lal</name></author></analytic><series><title level="j">Virology</title><idno type="ISSN">0042-6822</idno><imprint><date when="2007" type="published">2007</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>Cell Line</term><term>Chlorocebus aethiops</term><term>Cloning, Molecular</term><term>Cytoplasm (chemistry)</term><term>Immunoprecipitation</term><term>Microscopy, Confocal</term><term>Microscopy, Fluorescence</term><term>Protein Binding</term><term>SARS Virus (metabolism)</term><term>Two-Hybrid System Techniques</term><term>Viral Nonstructural Proteins (genetics)</term><term>Viral Nonstructural Proteins (metabolism)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Animaux</term><term>Clonage moléculaire</term><term>Cytoplasme ()</term><term>Immunoprécipitation</term><term>Liaison aux protéines</term><term>Lignée cellulaire</term><term>Microscopie confocale</term><term>Microscopie de fluorescence</term><term>Protéines virales non structurales (génétique)</term><term>Protéines virales non structurales (métabolisme)</term><term>Techniques de double hybride</term><term>Virus du SRAS (métabolisme)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Viral Nonstructural Proteins</term></keywords><keywords scheme="MESH" qualifier="chemistry" xml:lang="en"><term>Cytoplasm</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Protéines virales non structurales</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>SARS Virus</term><term>Viral Nonstructural Proteins</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Protéines virales non structurales</term><term>Virus du SRAS</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Cell Line</term><term>Chlorocebus aethiops</term><term>Cloning, Molecular</term><term>Immunoprecipitation</term><term>Microscopy, Confocal</term><term>Microscopy, Fluorescence</term><term>Protein Binding</term><term>Two-Hybrid System Techniques</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Animaux</term><term>Clonage moléculaire</term><term>Cytoplasme</term><term>Immunoprécipitation</term><term>Liaison aux protéines</term><term>Lignée cellulaire</term><term>Microscopie confocale</term><term>Microscopie de fluorescence</term><term>Techniques de double hybride</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) caused a severe outbreak in several regions of the world in 2003. The SARS-CoV genome is predicted to contain 14 functional open reading frames (ORFs). The first ORF (1a and 1b) encodes a large polyprotein that is cleaved into nonstructural proteins (nsp). The other ORFs encode for four structural proteins (spike, membrane, nucleocapsid and envelope) as well as eight SARS-CoV-specific accessory proteins (3a, 3b, 6, 7a, 7b, 8a, 8b and 9b). In this report we have cloned the predicted nsp8 gene and the ORF6 gene of the SARS-CoV and studied their abilities to interact with each other. We expressed the two proteins as fusion proteins in the yeast two-hybrid system to demonstrate protein-protein interactions and tested the same using a yeast genetic cross. Further the strength of the interaction was measured by challenging growth of the positive interaction clones on increasing gradients of 2-amino trizole. The interaction was then verified by expressing both proteins separately in-vitro in a coupled-transcription translation system and by coimmunoprecipitation in mammalian cells. Finally, colocalization experiments were performed in SARS-CoV infected Vero E6 mammalian cells to confirm the nsp8-ORF6 interaction. To the best of our knowledge, this is the first report of the interaction between a SARS-CoV accessory protein and nsp8 and our findings suggest that ORF6 protein may play a role in virus replication.</div></front></TEI><affiliations><list><country><li>Inde</li></country></list><tree><noCountry><name sortKey="Birch, Chris" sort="Birch, Chris" uniqKey="Birch C" first="Chris" last="Birch">Chris Birch</name><name sortKey="Catton, Mike" sort="Catton, Mike" uniqKey="Catton M" first="Mike" last="Catton">Mike Catton</name><name sortKey="Chow, Vincent T K" sort="Chow, Vincent T K" uniqKey="Chow V" first="Vincent T K" last="Chow">Vincent T K. Chow</name><name sortKey="Druce, Julian" sort="Druce, Julian" uniqKey="Druce J" first="Julian" last="Druce">Julian Druce</name><name sortKey="Fielding, Burtram C" sort="Fielding, Burtram C" uniqKey="Fielding B" first="Burtram C" last="Fielding">Burtram C. Fielding</name><name sortKey="Gunalan, Vithiagaran" sort="Gunalan, Vithiagaran" uniqKey="Gunalan V" first="Vithiagaran" last="Gunalan">Vithiagaran Gunalan</name><name sortKey="Lal, Sunil K" sort="Lal, Sunil K" uniqKey="Lal S" first="Sunil K" last="Lal">Sunil K. Lal</name><name sortKey="Liu, Boping" sort="Liu, Boping" uniqKey="Liu B" first="Boping" last="Liu">Boping Liu</name><name sortKey="Tan, Yee Joo" sort="Tan, Yee Joo" uniqKey="Tan Y" first="Yee-Joo" last="Tan">Yee-Joo Tan</name></noCountry><country name="Inde"><noRegion><name sortKey="Kumar, Purnima" sort="Kumar, Purnima" uniqKey="Kumar P" first="Purnima" last="Kumar">Purnima Kumar</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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