Stimulation by reticulocyte initiation factors of protein synthesis in a cell-free system from Krebs II ascites cells

Abstract

1. We have studied the effect of a crude initiation factor preparation, the 0.5 M KCl extract of reticulocyte ribosomes, on the translation of messenger RNAs in a cell-free system from Krebs II ascites cells. 2. Addition of these factors promotes the translation of lens and globin mRNAs and also stimulates the translation of a number of endogenous messengers. 3. Translation of encephalomyocarditis virus (EMC-) RNA is stimulated when protein synthesis is performed at high K + concentrations. At lower K + levels there is no effect or a slight inhibition. Poly(U)-directed incorporation is unaffected. 4. The reticulocyte factors enhance the incorporation of formylmethionine (fMet) from fMet-tRNA Met f into protein, and also the formation of fMet-puromycin, showing that their action is at the level of initiation. Results obtained with pactamycin and double-stranded RNA, both of which are inhibitors of initiation, confirm this conclusion. 5. The reticulocyte factor preparation also contains small amounts of globin mRNA which directs the synthesis of globin containing an excess of α chains. This mRNA can be inactivated by pancreatic ribonuclease without loss of the stimulatory activities of the initiation factors themselves.