In vitro synthesis of yeast ribosomal proteins

Abstract

Most yeast messenger RNAs contain a polyA fragment of about 50 nucleotides [l-4]. However, since a substantial amount of the pulse-labelled RNA from yeast polysomes lacks polyA [2,4] the presence of a polyA tract is not a property common to all yeast mRNAs. It is not known whether lack of polyA is limited to mRNA species coding for specific classes of protein or whether it is a reflection of the physiological state (e.g. age) of mRNA species in general. The only eukaryotic messenger RNA known to be devoid of polyA is histone messenger [5] . If indeed only specific mRNA species lack polyA, then messenger RNA for ribosomal protein, because of the physiological analogy between histones and ribosomal protein, would be a likely constituent of the mRNA fraction lacking polyA. Just as histones, ribosomal proteins are synthesized in the cytoplasm [6-81 and then transported into the nucleus [9]. The complex set of nucleocytoplasmic interactions required in each case might be influenced by the absence of polyA from the mRNA involved. If ribosomal protein messengers could be shown to occur in the fraction lacking polyA, isolation of these messengers and study of the genetic organization of the genes coding for ribosomal protein would be enormously facilitated. We separated polyA-containing mRNA from mRNA lacking polyA by chromatography on oligodTcellulose columns and translated both fractions in vitro using a wheat germ system. The products were analyzed using the method recently developed by us for identification of yeast ribosomal protein [lo]. The results presented in this paper demonstrate