Multiple glycoproteins synthesized by the smallest RNA segment (S10) of bluetongue virus

• Published in: Journal of Virology 1992-12, Vol.66 (12), p.7104-7112
• Main Authors: WU, X, SI-YI CHEN, IWATA, H, COMPANS, R. W, ROY, P
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; aparato golgi; appareil de golgi; arn; Base Sequence; Biological and medical sciences; bluetongue virus; Bluetongue virus - genetics; Bluetongue virus - metabolism; Cell Line; Codon - genetics; Cricetinae; DNA, Viral - genetics; Electrophoresis, Polyacrylamide Gel; endoplasmic reticulum; Fundamental and applied biological sciences. Psychology; glicoproteinas; glycoproteine; glycoproteins; Glycoproteins - biosynthesis; Glycoproteins - genetics; Glycoproteins - isolation & purification; golgi apparatus; Kidney; Microbiology; Molecular Sequence Data; Morphology, structure, chemical composition, physicochemical properties; Oligodeoxyribonucleotides; orbivirus; Plasmids; Polymerase Chain Reaction - methods; Restriction Mapping; reticulo endoplasmatico; reticulum endoplasmique; rna; RNA, Double-Stranded - genetics; RNA, Double-Stranded - metabolism; RNA, Viral - genetics; RNA, Viral - metabolism; Sequence Homology, Nucleic Acid; Viral Nonstructural Proteins - biosynthesis; Viral Nonstructural Proteins - genetics; Viral Nonstructural Proteins - isolation & purification; Virology
• ISSN: 0022-538X; 1098-5514
• DOI: 10.1128/jvi.66.12.7104-7112.1992

The genome of bluetongue virus, an orbivirus, consists of 10 double-stranded RNAs, each encoding at least one polypeptide. The smallest RNA segment (S10) encodes two minor nonstructural proteins, NS3 and NS3A, the structures and functions of which are not understood. We have expressed these two proteins in mammalian cells by using the T7 cytoplasmic transient expression system. Using a deletion mutant (lacking the first AUG initiation codon), we have demonstrated that the second initiation codon is used to initiate the synthesis of NS3A protein and that the two initiation codons are responsible for the synthesis not only of NS3 and NS3A but also of high-molecular-weight forms of both proteins. These higher-molecular-weight forms (GNS3 and GNS3A) are glycosylated. We have also demonstrated that the carbohydrate chains of GNS3 and GNS3A could be further modified by heterogeneous extension to polylactosaminoglycan forms. The glycosylated and nonglycosylated forms are found in similar intracellular locations in the Golgi complex. In the presence of cycloheximide, NS3 and NS3A immunofluorescence staining was pronounced in the Golgi complex, confirming that NS3 and NS3A are competent for transport to the Golgi apparatus after synthesis. We conclude that S10 gene products are integral membrane glycoproteins.