Further studies on nuclear ribonucleoprotein particles containing DNA-like RNA from rat liver

Abstract

Nuclear ribonucleoprotein particles containing DNA-like RNA have been extracted from purified rat liver nuclei in the presence of high levels of cytoplasmic ribonuclease inhibitor. The following facts have been noted: 1. 1. After labelling with radioactive orotic acid, nuclear ribonucleoprotein particles sedimenting between 30 and 400 S with a maximum distribution at 180 S have been isolated. Pulse labelling of the RNA with pulses of up to 40 sec leads to the isolation of ribonucleoprotein particles sedimenting mainly in the light region of the sucrose gradient at 50–200 S. Inhibition of RNA synthesis by actinomycin D or α-amanitin also causes a reduction of the sedimentation coefficients of the labelled nuclear particles. 2. 2. Independent of their size, the ribonucleoprotein particles have a narrow bouyant density distribution at 1.39–1.40 g/cm 3 corresponding to a protein: RNA ratio of 4 : 1. As shown by dodecyl sulfate-polyacrylamide gel electrophoresis, the protein composition of the ribonucleoprotein particles is the same independent of the sedimentation coefficients. The pattern of band distribution indicates a considerable heterogeneity of proteins. The molecular weights were calculated to be 32 000 for the lightest and 150 000–200 000 for the heaviest polypeptide chain. 3. 3. The RNA isolated from the ribonucleoprotein particles is highly heterogeneous and sediments between 5 S and 80 S. The sedimentation behavior of the RNA is correlated to the size of ribonucleoprotein particles from which the RNA had been extracted.