Synthesis of Vimentin in a Reticulocyte Cell‐Free System Programmed by Poly(A)‐Rich RNA from Several Cell Lines and Rat Liver

Abstract

Poly(A)‐rich RNA has been isolated from Ehrlich ascites tumour (EAT) cells and translated in a rabbit reticulocyte cell‐free system. The intermediate filament protein, vimentin, was found to be a major translation product. Fractionation of the poly(A)‐rich RNA by sucrose gradient centrifugation showed that the vimentin mRNA had a sedimentation coefficient of about 18 S corresponding to a molecular size of about 2000 nucleotides. This means that it must possess significant non‐coding regions. Vimentin synthesized in vitro was identical to native vimentin with regard to its precipitability with ammonium sulphate, extent of phosphorylation and susceptibility to digestion by the vimentin‐specific, Ca²⁺‐activated proteinase. Poly(A)‐rich RNA was also isolated from a number of tissue‐culture cells and rat liver, which contain varying amounts of vimentin in situ. It was found that the amount of vimentin synthesized by these RNA preparations in a rabbit reticulocyte cell‐free system is proportional to the amount of vimentin detectable in situ, suggesting that the amount of cellular vimentin may be controlled at the level of transcription.