Nucleocytoplasmic translocation of RNA in Tetrahymena pyriformis and its inhibition by actinomycin D and cycloheximide

Abstract

The nucleocytoplasmic translocation of RNA pulse-labelled with 3H-uridine was studied in logarithmically growing Tetrahymena pyriformis GL cells during a subsequent chase period, without or with actinomycin (AMD) or cycloheximide (CH), by light and electron microscopic autoradiography and analysis of RNA by electrophoresis, and in isolated cell fractions (nuclei, nuclear membranes, nuclear non-membrane bulk material, microsomes, and post-microsomal fractions). Kinetic changes in nucleocytoplasmic transfer of RNA as well as changes of RNA contents and fine structure brought about by AMD and CH are described. Both antibiotics applied at concentrations inhibitory to RNA synthesis effectively inhibited the nucleocytoplasmic translocation of pre-existing nuclear RNA and resulted in relative accumulation of stable RNA moieties, especially of precursors of rRNA, in the macronucleus. The marked stability of the apparent primary transcription product of the nucleoli (about 2.2 million D mol. wt) indicated that neither considerable processing nor degradation took place during 90 min of chase in the presence of both drags. RNA with the molecular weights of mature rRNA (1.32 and 0.7 million D) was not found in substantial amounts in purified macronuclei from normal cells but was detected in the nuclei from AMD-treated cells. AMD completely inhibited nucleocytoplasmic migration of pulse-labelled RNA, the onset of this event being correlated with the time needed for a complete inhibition of 3H-uridine incorporation at a particular drag concentration. With CH, however, even when applied at relatively high concentrations, a residual translocation amounting to 15–20% of that of controls was maintained throughout the chase period.