Expression of the RNA genome of an animal virus in Saccharomyces cerevisiae.
The nucleocapsid of vesicular stomatitis virus (VSV) was introduced into the cytoplasm of Saccharomyces cerevisiae by low pH-dependent fusion of the viral envelope with the spheroplast plasma membrane. This led to de novo synthesis of the three major structural proteins of the virus--the G, N, and M proteins--as shown by immunoprecipitation of [35S]methionine-labeled spheroplast lysates. In NaDodSO4/polyacrylamide gel electrophoresis, M and N proteins comigrated with those of the virion, whereas the yeast-made G protein migrated as two bands with apparent molecular sizes of 60 and 70 kDa. Both polypeptides appeared to be N-glycosylated, since only one polypeptide with the apparent molecular mass of approximately equal to 55 kDa was produced in the presence of tunicamycin. Phase separation into Triton X-114 suggested that the unglycosylated G protein was membrane bound. According to immunofluorescent surface staining of live spheroplasts, at least part of the G protein was transported to the plasma membrane. Spheroplasts expressing the VSV genes could be fused together by low pH to form polykaryons, indicating that G protein synthetized by yeast was fusogenic--i.e., biologically active.
D000818:Animals; D001692:Biological Transport; D002213:Capsid / Q000096:biosynthesis; D005814:Genes, Viral; D006023:Glycoproteins / Q000378:metabolism; D008562:Membrane Glycoproteins; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics*; D014758:Viral Core Proteins / Q000096:biosynthesis; D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins / Q000096:biosynthesis* / Q000378:metabolism; D015678:Viral Structural Proteins
C028391:G protein, vesicular stomatitis virus; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D014758:Viral Core Proteins; D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins; D015678:Viral Structural Proteins