Molecular biology of the Severe Acute Respiratory Syndrome coronavirus (SARS -CoV) accessory protein...

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" Molecular biology of the Severe Acute Respiratory Syndrome coronavirus (SARS -CoV) accessory proteins ORF7a and ORF7b "
Scott Richard Schaecher A. Pekosz

Document Type	:	Latin Dissertation
Language of Document	:	English
Record Number	:	54461
Doc. No	:	TL24415
Call number	:	‭3316676‬
Main Entry	:	Scott Richard Schaecher
Title & Author	:	Molecular biology of the Severe Acute Respiratory Syndrome coronavirus (SARS -CoV) accessory proteins ORF7a and ORF7b\ Scott Richard Schaecher
College	:	Washington University in St. Louis
Date	:	2008
Degree	:	Ph.D.
student score	:	2008
Page No	:	295
Abstract	:	The Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) was identified as the etiologic agent of the first global pandemic of the 21st century. Like other viruses within the Coronaviridae, SARS-CoV expresses canonical replication and structural genes as well as a highly divergent set of accessory genes whose open reading frames are interspersed among the structural genes. Annotation of the SARS-CoV genome predicted eight accessory genes, the most of any known coronavirus. The primary aim of this thesis was to characterize the molecular biology of the two accessory proteins expressed from viral subgenomic RNA number 7 (sgRNA 7), ORF7a and ORF7b. ORF7a is an integral membrane protein of unknown function that localizes to the Golgi apparatus. Expression of the putative ORF7b protein in virus infected cells was confirmed and mutational analysis of gene 7 cDNA altering the translational contexts of both ORF7a and ORF7b revealed that translation initiation of ORF7b occurs via ribosomal leaky scanning. Biochemical characterization revealed that ORF7b is likely a type III integral membrane protein, it localizes exclusively to the Golgi region in transfected and infected cells, and the ORF7b transmembrane domain is necessary and sufficient for Golgi retention. Electron microscopy and biochemical analysis has revealed that the ORF7b protein is packaged into virus particles, suggesting a potential role in virus assembly, entry, or uncoating. Both ORF7a and ORF7b likely provide "luxury" functions for the virus, as gene 7 deletion viruses replicated to wild-type levels in vitro. Although the deletion viruses also induced early stages of apoptosis similar to wild-type SARS-CoV, the mutant viruses triggered an altered apoptotic cascade resulting in significant reductions in oligonucleosomal DNA fragmentation. To determine contributions of the gene 7 proteins to virus replication and pathogenesis in vivo, an immunocompromised Syrian golden hamster model for SARS-CoV infection was developed. Utilizing cyclophosphamide to induce a transient immunosuppression resulted in severe morbidity and mortality among hamsters infected with SARS-CoV. Although the gene 7 deletion viruses replicated to wild-type levels, had no altered tropism, and resulted in no significant difference in pathogenesis or mortality compared to wild-type virus, this small animal model may be of significant use in analyzing other viral accessory genes as well as screening of antiviral compounds.
Subject	:	Biological sciences; Accessory proteins; Coronavirus; SARS-CoV; Severe acute respiratory syndrome; Molecular biology; Virology; 0720:Virology; 0307:Molecular biology
Added Entry	:	A. Pekosz
Added Entry	:	Washington University in St. Louis