Nuclear RNA fractions obtained by sequential extraction of rat liver nuclei with salt solutions and their relationship to polysome-associated RNA.

Abstract

Pulse‐labeled RNA from rat liver nuclei was fractionated by sequential treatment of purified nuclei with buffered salt solutions of different ionic strength in the presence of ribonuclease inhibitor prepared from rat liver cytoplasm. Two fractions of pulse‐labeled nuclear RNA were successively extracted from rat liver nuclei with 0.14 M NaCl, 0.001 M MgCl2 at pH 7.0 and pH 8.0, respectively. The labeled material isolated at pH 7.0 sediments between 6–16 S while the pulse‐labeled RNA extracted at pH 8.0 sediments in the range of 10–34 S. Both fractions are characterized by a high content in AMP. With respect to [32P]nucleotide composition, sedimentation characteristics and hybridizability, these fractions correspond closely to non‐ribosomal RNA associated with polysomes. When a third extraction is carried out at pH 8.0 in 0.3 M NaCl another rapidly labeled RNA component is obtained. This RNA sediments above 34 S, turns over more rapidly than the RNA species extracted in 0.14 M NaCl, and contains less AMP. As judged by [32P]base‐composition, sedimentation and hybridizability, this RNA component is not present in polysomes. The pulse‐labeled RNA remaining in the nuclei after three salt extractions comprises mainly ribosomal or ribosomal precursor RNA as well as DNA‐like RNA species which show little or no nucleotide sequence homology with polysome‐associated RNA.Since nuclear RNA species clearly different from polysome‐associated cytoplasmic RNA are released from nuclei only at a relatively high ionic strength and are retained at a physiologic salt concentration it is suggested that RNA species not designated to function in cytoplasmic protein synthesis are a priori bound more tightly to nuclear structures than those types of RNA which serve as vehicles for the nucleo‐cytoplasmic transfer of genetic information.