Preliminary characterization of molecules that increase cell free translational activity of cardiac cytoplasmic RNA

Previous evidence indicates that cells generate molecules that alter transcription in response to stress. Accordingly, we conducted preliminary experiments to characterize these molecules. Extracts were prepared from normal and experimental canine hearts in which a 100 mmHg gradient was created across the ascending aorta for 6 h. Experimental extracts were then treated by ultrafiltration through YM 10 and YM 30 membranes, by incubation with immobilized trypsin for 1 h at 37 degrees C and by incubation in a boiling water bath for 10 min. Extracts were perfused through isolated rat hearts for 1 h. Total cytoplasmic RNA was then extracted from the perfused heart and translated in a cell free medium containing [35S]-methionine. Incorporated label into newly synthesized protein was determined by liquid scintillation counting. A 13% mean increase in translational activity was produced by the fraction of the experimental extract passing through the YM 10 membrane compared with the material retained by YM 10 and YM 30 membranes. A 14% mean decrease in translational activity was observed in hearts perfused with experimental extracts treated with immobilized trypsin compared to untreated experimental extracts. There was no significant difference in translational activity of hearts perfused with experimental extracts subjected to boiling compared with non-boiled experimental extracts. These data suggest that the active molecules may be heat stable peptides or peptide containing substances of 10000 daltons or less in molecular weight.