Translational regulation of the synthesis of a major heat shock protein in HeLa cells.
The synthesis of a major heat shock protein (HSP 70) was measured in HeLa cells incubated at 42.5 degrees C and then transferred to 37 degrees C or 30 degrees C. After 90 min, synthesis of HSP 70 decreased by 54 and 85%, respectively, whereas HSP 70 mRNA was reduced at most by 20%. Therefore, the reduced synthesis of HSP 70 could not be accounted for by mRNA turnover. HSP 70 was associated with large polyribosomes (6-10 ribosomes) in cells kept at 42.5 degrees C, but with medium or small polyribosomes in cells transferred to 37 degrees C or 30 degrees C (5-6 or 2-3 ribosomes, respectively). Addition of puromycin to these cells resulted in the release of all ribosomes from HSP 70 mRNA, indicating that they were translationally active. The regulation of HSP 70 synthesis was investigated in cell-free systems prepared from heat-shocked or control cells and incubated at 30 degrees C and 42 degrees C. After 5 min at 42 degrees C, the cell-free system from heat-shocked cells synthesized protein at 3 times the rate of the control cell-free system. This difference was in large part due to synthesis of HSP 70. Addition of HSP mRNA to the control cell-free system stimulated protein synthesis at 42 degrees C, but not at 30 degrees C. These findings suggest that translation of HSP 70 mRNA is specifically promoted at high temperature and repressed during recovery from heat shock by regulatory mechanisms active at the level of initiation.
D002474:Cell-Free System; D003609:Dactinomycin / Q000494:pharmacology; D006367:HeLa Cells; D006360:Heat-Shock Proteins / Q000096:biosynthesis* / Q000235:genetics; D006358:Hot Temperature; D006801:Humans; D007700:Kinetics; D011132:Polyribosomes / Q000378:metabolism; D014176:Protein Biosynthesis; D011691:Puromycin / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism
D006360:Heat-Shock Proteins; D012333:RNA, Messenger; D003609:Dactinomycin; D011691:Puromycin