Transport of β‐globin mRNA from nuclei of murine friend erythroleukemia cells

Abstract

An in vitro assay system for analysis of .beta.-globin mRNA transport is described. Nuclei isolated from murine Friend erythroleukemia cells induced to synthesize globin mRNA, were incubated in microassays. By electrophoresis and hybridization analysis, released 9-S .beta.-globin mRNA was shown to be undegraded. After direct blotting, the released mRNA was quantified by hybridization with a labeled plasmid containing a .beta.-globin DNA restriction fragment. The inducibility of .beta.-globin mRNA transport corresponded to that previously reported for the release of rapidly labeled RNA in other assay systems. In contrast to the ineffectiveness of high concentrations of the SH reagent iodoacetate, low concentrations of the oxidizing SH reagent, o-iodosobenzoate, inhibited the release of .beta.-globin mRNA from nuclei of erythroleukemia cells, as well as the release of rapidly labeled RNA from rat liver nuclei. The inhibitory effect of the oxidizing agent on .beta.-globin mRNA transport could be reversed by postincubation of the nuclei with the reducing agent, dithiothreitol. The potential role of disulfide bond formation on RNA transport is discussed.