Effect of extraction temperature on template activity of polysomal RNA from fetal, neonatal, and adult rat liver

Abstract

Polyribosomes were prepared from adult rat liver and the RNA was extracted from the polysomes at various temperatures. The ability of these RNA fractions to serve as templates in the polymerization of amino acids, i.e., mRNA, was tested in a heterologous cell-free system from E. coli D 10, a mutant which is devoid of RNase I. Electrophoresis of the adult liver polysomal RNA extracted at 37, 45, 55, 65, and 70 ° on polyacrylamide gels and fractionation on linear sucrose gradients indicated the presence of species of RNA with molecular weights in excess of 28S ribosomal RNA. This represented aggregation of the latter. The highest template activity was associated with the RNA extracted at 55 °. In this regard, the 28S region of the sucrose gradient was the most active although activity was seen throughout most of the gradient. Polyribosomes were also prepared from fetal and neonatal rat liver; the RNA was extracted at 55 °. Both fetal and neonatal rat liver polysomal RNA exhibited at least a 2-fold greater total template activity than adult material. The template activity of neonatal and fetal RNA fractions from sucrose gradients had very similar patterns although fetal RNA was slightly more efficacious. With these fractions, highest total template activity was also seen in the 28S region of the gradients. The maximal specific template activity, i.e., amount labeled amino acid incorporated per μg RNA, however, occurred in the region between 6 and 18S RNA.