Isolation and visualization of large compact ribonucleoprotein particles of specific nuclear RNAs

Abstract

We have previously shown that nuclear transcripts of the multifunctional enzyme, carbamoyl-phosphate synthetase, aspartate transcarbamylase, dihydroorotase RNA can be released from nuclei of Syrian hamster cells as compact ribonucleoprotein (RNP) particles that sediment at the 200S region in a sucrose gradient. The 200S nuclear RNP particles contain U1, U2, and U6 small nuclear RNPs, which are known to be required for splicing of pre-mRNA, as integral components of the particles. In this study we demonstrate that nuclear transcripts of dihydrofolate reductase in Syrian hamster cells and of beta-actin in both Syrian hamster and human cells are also released from the respective nuclei as 200S particles--despite the difference in length of these RNAs. Electron microscopy of the 200S particles revealed discrete compact composite structures with a cross section of approximately 50 nm. Finding that two more nuclear RNAs from two different cell types and two different species are released as 200S RNP particles suggests a general mode for packaging of heterogeneous nuclear RNA in large compact RNP particles the size of which is independent of the RNA length.