Genetic and Molecular Biological Analysis of Protein-Protein Interactions in Coronavirus Assembly

New York State Department of Health, Albany, New York, USA.
Advances in Experimental Medicine and Biology (Impact Factor: 1.96). 02/2006; 581:163-73. DOI: 10.1007/978-0-387-33012-9_29
Source: PubMed


Virions of coronaviruses (CoVs) are pleiomorphic, with a roughly spherical structure brought about by cooperation among a relatively small set of structural proteins and a membranous envelope acquired from the endoplasmic reticulum–Golgi intermediate compartment (ERGIC) (Fig. 1). Three integral membrane proteins reside in the envelope. The most salient of these is the spike glycoprotein (S), which mediates receptor attachment and fusion of the viral and host cell membranes. The membrane protein (M) is the most abundant virion component and gives the envelope its shape. The third constituent is the envelope protein (E), which, although minor in both size and quantity, plays a decisive role is envelope formation. In some group 2 CoVs, an additional protein, the hemagglutinin-esterase (HE), appears in the viral envelope. Finally, interior to the envelope, monomers of the nucleocapsid protein (N) wrap the genome into a helical structure.

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    • "Synthesis of a 3′ co-terminal nested set of sgmRNAs is a common feature of members of the Nidovirales [9], [15], [52]–[53]. In addition to this feature, the 5′ end of all sgmRNAs in arteri- and coronaviruses possesses a common leader sequence derived from the 5′ terminus of the genomic RNA [2], [7]–[9], [15], [17], [54]. "
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    ABSTRACT: The establishment of persistent viral infection is often associated with the selection of one or more mutant viruses. For example, it has been found that an intraleader open reading frame (ORF) in genomic and subgenomic mRNA (sgmRNA) molecules is selected during bovine coronavirus (BCoV) persistence which leads to translation attenuation of the downstream ORF. Here, we report the unexpected identification of leaderless genomes, in addition to leader-containing genomes, in a cell culture persistently infected with BCoV. The discovery was made by using a head-to-tail ligation method that examines genomic 5'-terminal sequences at different times postinfection. Functional analyses of the leaderless genomic RNA in a BCoV defective interfering (DI) RNA revealed that (1) the leaderless genome was able to serve as a template for the synthesis of negative-strand genome, although it cannot perform replicative positive-strand genomic RNA synthesis, and (2) the leaderless genome retained its function in translation and transcription, although the efficiency of these processes was impaired. Therefore, this previously unidentified leaderless genome is associated with the attenuation of genome expression. Whether the leaderless genome contributes to the establishment of persistent infection remains to be determined.
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  • No preview · Article · Feb 2006 · Advances in Experimental Medicine and Biology
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    ABSTRACT: A deletion mutant of severe acute respiratory syndrome coronavirus (SARS-CoV) has been engineered by deleting the structural E gene in an infectious cDNA clone that was constructed as a bacterial artificial chromosome (BAC). The recombinant virus lacking the E gene (rSARS-CoV-DeltaE) was rescued in Vero E6 cells. The recovered deletion mutant grew in Vero E6, Huh-7, and CaCo-2 cells to titers 20-, 200-, and 200-fold lower than the recombinant wild-type virus, respectively, indicating that although the E protein has an effect on growth, it is not essential for virus replication. No differences in virion stability under a wide range of pH and temperature were detected between the deletion mutant and recombinant wild-type viruses. Although both viruses showed the same morphology by electron microscopy, the process of morphogenesis seemed to be less efficient with the defective virus than with the recombinant wild-type one. The rSARS-CoV-DeltaE virus replicated to titers 100- to 1,000-fold lower than the recombinant wild-type virus in the upper and lower respiratory tract of hamsters, and the lower viral load was accompanied by less inflammation in the lungs of hamsters infected with rSARS-CoV-DeltaE virus than with the recombinant wild-type virus. Therefore, the SARS-CoV that lacks the E gene is attenuated in hamsters, might be a safer research tool, and may be a good candidate for the development of a live attenuated SARS-CoV vaccine.
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