Biological activity of 125iodine labelled globin mRNA in an Ehrlich ascites cell-free system

Globin mRNA was isolated from avian erythroblasts and labelled with ¹²⁵iodine to a specific activity of 4.5×10⁶ dpm μg^-1 mRNA. The labelling procedure was modified as compared to common techniques in order to protect the mRNA against degradation. When this RNA is translated in an Ehrlich ascites cell-free system globin chains are synthesized as is demonstrated by the analysis of the in vitro synthesized products. The biological activity of ¹²⁵iodine labelled mRNA has proven to be 30% of that of unlabelled mRNA.     

Translation of pure feather keratin mRNA in a wheat embryo cell-free system

Highly purified feather keratin mRNA, prepared by dissociation of mRNP particles in Na dodecyl sulphate, was translated in a wheat embryo cell-free system with similar efficiency to rabbit globin mRNA and RNA purified from cucumber mosaic virus. The only detectable products of translation of the keratin mRNA were keratin chains, which were identical to native keratin chains as judged by several different criteria. These results support previous conclusions that the keratin mRNA can be obtained in a pure state.     

Translation of hepatic mRNA in extracts from wheat germ embryos

Extracts from wheat embryos have been used to study the incorporation of amino acids into TCA insoluble products using hepatic mRNA fractions. The properties of this system are described and compared to the incorporation obtained with poly U and rabbit globin mRNA. SDS-acrylamide gel analysis showed that the major polypeptide synthesized with globin mRNA co-migrates with rabbit globin (15 500 daltons). Rat liver products were numerous, with molecular weights from less than 10000 to greater than 65000 daltons. The KCl concentration for maximum incorporation into TCA precipitable polypeptides with hepatic mRNA was not the optimum KCl concentration for synthesis of complete products.     

Effects of cordycepin,hydroxyurea and cycloheximide on histone mRNA synthesis in synchronized HeLa cells

The effects of cordycepin, hydroxyurea and cycloheximide on the synthesis of histone mRNA in synchronized HeLa cells were studied by quantitating the RNA, using translation in a rabbit reticulocyte cell-free system. It was found that biologically active histone mRNA is synthesized in the presence of cordycepin. This result strengthens the findings by others that histone mRNA does not contain poly(A). Hydroxyurea and cycloheximide, when used at concentrations that inhibit cellular DNA synthesis, had different effects on histone messenger activity. While polyribosomal messenger activity rapidly declined after addition of hydroxyurea it was not impaired by cycloheximide. These results might help to elucidate regulatory mechanisms involved in the coupled synthesis of DNA and histones in HeLa cells.