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Organization of the M genomic segment of Toscana phlebovirus
P Di Bonito1, S Mochi, M C Grò
1Laboratory of Virology, Istituto Superiore di Sanità, Rome, Italy.
Abstract:
The nucleotide sequence of the Toscana (TOS) virus M RNA segment contains a single major open reading frame in the viral-complementary sequence, which can encode a polyprotein of 1339 amino acids. To map the TOS M segment product(s), different regions of the putative M polypeptide were expressed as glutathione S-transferase fusion proteins, which were purified and inoculated into mice to produce hyperimmune sera. By Western blot analysis, a protein of approximately 30 kDa and two glycoproteins, G1 and G2, with the same molecular mass (approximately 65 kDa) were identified in TOS virus-infected cells. The 30 kDa protein, which reacted with antibodies raised to the NH2-terminal, was found to be a non-structural protein (designated NSm). By immunoprecipitation analysis of TOS virus-infected cell lysates, both treated or untreated with tunicamycin, the relative positions of glycoproteins G1 and G2 were determined. The gene order, with respect to the genomic M RNA, was found to be 3' NSm-G1 -G2 5'
Insights
Researchers identified Toscana virus M segment proteins, including a non-structural protein (NSm) and two glycoproteins (G1, G2). This study clarifies the gene order of the Toscana virus M RNA segment.
Area of Science:
- Virology
- Molecular Biology
- Protein Characterization
Background:
- The Toscana (TOS) virus M RNA segment encodes a large polyprotein.
- Understanding the TOS virus M segment's protein products is crucial for viral research.
Purpose of the Study:
- To map the protein products encoded by the Toscana virus M RNA segment.
- To determine the gene order of the TOS virus M RNA.
Main Methods:
- Expression of glutathione S-transferase fusion proteins from TOS M segment regions.
- Production of hyperimmune sera in mice.
- Western blot and immunoprecipitation analyses of TOS virus-infected cells.
Main Results:
- Identified a 30 kDa non-structural protein (NSm) and two glycoproteins (G1, G2) of approximately 65 kDa in TOS virus-infected cells.
- Determined the gene order on the M RNA segment as 3' NSm-G1-G2 5'.
Conclusions:
- The TOS virus M RNA segment produces a non-structural protein (NSm) and two glycoproteins (G1, G2).
- The established gene order provides a foundational understanding of TOS virus M segment gene expression.