Differences in the primary structure of nucleolar and ribosomal high molecular weight RNA of the novikoff hepatoma and rat liver, and possible therapeutic implications.

Abstract

The remarkable advances in the methods for the isolation of nuclei and nucleoli of cancer cells along with the rapid evolution of techniques for purification and structural analyses of high molecular weight RNA has provided new opportunities to comparatively evaluate the constituents of tumor cells and other cells. Complete pancreatic RNase digestion was carried out on highly purified nucleolar 45S, 35S and 28S RNA and ribosomal 28S and 18S RNA of Novikoff hepatoma ascites cells which were highly labeled in vitro with 32P-orthophosphate. In addition, rat liver ribosomal 28S RNA was labeled in vivo and subjected to the same isolation and RNase digestion procedure. The resultant oligonucleotides of the digests are purine containing products with a pyrimidine 3′ terminal, altogether referred to as the pyrimidine catalog of the RNA molecule. They were separated on DEAE Sephadex A-25 at pH 7.6 according to chain length and the single components were isolated by paper electrophoresis at pH 3.5 (mono- and dinucleotides) or by rechromatography on DEAE Sephadex A-25 at pH 3.3 (hepta- to tridecanucleotides). Sequence studies were carried out by means of nucleotide analysis, complete digestion with T1 RNase and partial digestion with spleen phosphodiesterase. The oligonucleotide sequences and their frequencies were compared between the different high molecular weight RNA species of the Novikoff hepatoma and 28S ribosomal RNA of rat liver. Interestingly, marked differences were found between the substructures of nucleolar 45S, 35S and 28S RNA compared to those of ribosomal 18S RNA. The longest polypurine sequences of 28S ribosomal RNA were identical to those of 28S nucleolar RNA and no differences were found between polypurines of the Novikoff hepatoma and rat liver. However, when the frequencies were determined for the mono- and dinucleotides, significant differences appeared mainly for the dinucleotide G-Up. On the basis of a calculated chain length of 4.500 for ribosomal 28S RNA, 134 G-Up residues were calculated for the Novikoff hepatoma compared to 103 for 28S rRNA of rat liver. These results provide further evidence for defined sequence differences between high molecular weight RNA of tumor cells and other cells and possibly point the way to definitive cancer chemotherapy. Critical evaluations of potential strategies are discussed.