Antibody-mediated polysome precipitation as a method for the size determination of viral mRNA species: viral envelope glycoprotein mRNA of avian sarcoma viruses.
A method was developed that allows the in situ isolation of viral mRNA, i.e. from polysomes of infected cells. This was achieved by precipitating intact polysomes via their nascent virus polypeptides using virus-specific antibodies. As a model, antibodies to the major envelope glycoprotein (gp85) of avian sarcoma viruses were employed to precipitate those polysomes from infected cells which synthesized the corresponding p70 virus glycoprotein precursor. Normal immunoglobulin and polysomes from uninfected chicken embryo fibroblasts served as controls. The radioactivity labelled mRNA from antibody-precipitated polysomes could subsequently be extracted and characterized for size. It was found that avian sarcoma virus gp85 envelope glycoprotein is predominantly synthesized by a 22--28 s viral mRNA. In addition, minor amounts of gp85-specific mRNAs of 16--21 s and 28--35 s could be demonstrated. The data indicate the presence in polysomes of viral mRNA species coding for i) gp85 only (16--21 s RNA), ii) gp85 and pp60src protein from the adjacent src-gene (22--28 s RNA), and iii) a large viral precursor protein (28--35 s RNA).
D000818:Animals; D000914:Antibodies, Viral; D001358:Avian Sarcoma Viruses / Q000032:analysis*; D002458:Cell Fractionation; D002460:Cell Line; D011232:Chemical Precipitation; D002642:Chick Embryo; D008722:Methods; D011132:Polyribosomes; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification*; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology
D000914:Antibodies, Viral; D012333:RNA, Messenger; D012367:RNA, Viral; D014759:Viral Envelope Proteins; D014764:Viral Proteins