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Identification and initial characterization of the murine gammaherpesvirus 68 gene M3, encoding an abundantly
V van Berkel1, K Preiter, H W Virgin
1Center for Immunology, and Departments of Pathology and Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri, USA.
Abstract:
Several viruses, including members of the gammaherpesvirus family, encode proteins that are secreted into the extracellular environment. We have identified an abundant 44-kDa secreted protein that is present in the supernatant of fibroblasts infected with murine gammaherpesvirus 68 (gammaHV68; also referred to as MHV-68) but not in that of uninfected fibroblasts. Sequence analysis of the amino terminus and of internal peptides revealed that this protein is encoded by the gammaHV68 M3 open reading frame (ORF). The amino-terminal sequence of the secreted protein starts at residue 25 of the M3 ORF, consistent with the first 24 residues functioning as a signal peptide. Northern blot analysis revealed a single abundant approximately 1.4-kb early-late lytic transcript encoded by the M3 ORF. Analysis of a partial cDNA clone and subsequent analyses of products of rapid amplification of cDNA ends coupled with S1 nuclease protection assays demonstrate that the M3 protein is encoded by an unspliced, polyadenylated mRNA initiating at bp 7294 and terminating at bp 6007 of the gammaHV68 genome. The 3' end of the M3 transcript maps 9 bp downstream of a consensus polyadenylation signal. Thus, the predicted M3 ORF is a functional gene that encodes an abundant secreted protein which is a candidate for interacting with host cellular receptors or cytokines.
Insights
Researchers identified a secreted protein from murine gammaherpesvirus 68 (gammaHV68) infection. This abundant protein, encoded by the M3 open reading frame (ORF), is secreted into the extracellular environment.
Area of Science:
- Virology
- Molecular Biology
- Immunology
Background:
- Gammaherpesviruses encode secreted proteins.
- Secreted viral proteins can interact with host factors.
Purpose of the Study:
- Identify and characterize secreted proteins from murine gammaherpesvirus 68 (gammaHV68).
- Determine the genetic origin and expression of a novel secreted protein.
Main Methods:
- Fibroblast infection assays with gammaHV68.
- Protein sequencing (amino terminus and internal peptides).
- Northern blot analysis.
- cDNA cloning and analysis (RACE, S1 nuclease protection assays).
Main Results:
- Identified an abundant 44-kDa secreted protein in gammaHV68-infected fibroblast supernatant.
- Protein is encoded by the M3 open reading frame (ORF) of gammaHV68.
- M3 ORF produces a single, abundant, unspliced, polyadenylated mRNA transcript.
- The first 24 residues of M3 ORF likely function as a signal peptide.
Conclusions:
- The M3 ORF is a functional gene encoding an abundant secreted protein.
- This secreted M3 protein is a potential mediator of host-virus interactions, possibly binding cellular receptors or cytokines.