Characterization of poly(A +)RNA in free messenger ribonucleoprotein and polysomes of mouse Taper ascites cells

Abstract

Cytoplasmic extracts of mouse Taper ascites cells were centrifuged on sucrose gradients to give 0–80 S, monosome, and polysome fractions. CsCl equilibrium density centrifugation of formaldehyde-fixed material from the 0–80 S fraction demonstrated that the messenger RNA in the 0–80 S fraction was in the form of free ribonucleoprotein. The size of the poly(A +)RNA and the size of the poly(A) segments of these molecules were shown to be very similar in both the free mRNP † † Abbreviation used:RNP,ribonucleoprotein. and polysome fractions. The labeling kinetics of the free mRNP poly(A +)RNA was similar to that of the polysomal poly(A +)RNA. The free mRNP poly(A +)RNA efficiently stimulated protein synthesis in the wheat germ cell-free system, supporting the view that it was mRNA. Two-dimensional gel electrophoresis was used to analyze the proteins whose synthesis was directed by free mRNP and polysomal poly(A +)RNA. The free mRNP poly(A +)RNA directed the synthesis of a simpler set of abundant protein products than did the polysomal poly(A +)RNA. Most of the free mRNP abundant protein products were also present in the polysomal products, though obvious quantitative differences were evident, indicating that each individual mRNA had its own characteristic distribution between polysomes and the translationally inactive RNP form.