Separation of mercury substituted RNA synthesized in isolated rat liver nuclei

Abstract

The population of RNA molecules synthesized in isolated rat liver nuclei in vitro in the presence of [3H]CTP and Hg-UTP was successfully fractionated into at least two subfractions containing various proportions of mercury label. Fractionation was achieved either by step-wise chromatography of Hg-RNA on thiopropyl-Sepharose columns or by density gradient centrifugation in metrizamide. The fraction of RNA heavily labeled with Hg-UTP was composed mainly of 4--18S RNA and contained virtually all radioactivity derived from [gamma-32P]ATP or [gamma-32P]GTP. The slightly mercurated RNA fraction consisted mainly of longer RNA molecules (12- greater than 28S) and was not labeled with [gamma-32P]ATP or [gamma-32P]GTP. Labeling with gamma-32P nucleoside triphosphates was sensitive both to rifamycin AF/013 and heparin whereas labeling with [3H]CTP was fully resistant to the inhibitors and showed sensitivity to low doses of alpha-amanitin. We assume that the observed subpopulation of heavily mercurated RNAs consists of RNA molecules initiated in vitro.