Control of globin synthesis by hemin Effect of temperature on globin synthesis by the reticulocyte cell-free system and the activity of translational repressors formed in the absence of hemin

Abstract

The relationship between temperature and the hemin dependence of globin synthesis by the rabbit reticulocyte cell-free system has been investigated. In the absence of hemin, globin synthesis almost ceases after 5 min at 34 °C but continues at a linear rate for 2 h at 25 °C. If, after 10 or 20 min of incubation at 34 °C in the absence of hemin, the reticulocyte cell-free system is cooled to 25 °C, globin synthesis resumes immediately at the rate of samples incubated at 25 °C from time zero. If the ribosome-free supernatant fraction is incubated without hemin at 25 °C, inhibitory activity which can be inactivated by hemin (reversible inhibitor) appears within 1 h and an inhibitory activity which is insensitive to hemin (irreversible inhibitor) is generated after a 2-h delay. These inhibitors are assayed by addition to fresh cell-free systems incubated at 34 °C. The reversible inhibitor, whether formed at 25 °C or 34 °C, has no effect on fresh cell-free systems when incubated at 25 °C without hemin. The irreversible inhibitor is less inhibitory at 25 °C than at 34 °C. The results suggest that globin synthesis is independent of hemin for 2 h at 25 °C because of the delay in formation of the irreversible inhibitor. Globin synthesis resumes when cell-free systems are first briefly incubated without hemin at 34 °C and then cooled to 25 °C, because the reversible inhibitor formed at 34 °C is ineffective at the lower temperature. If the reversible inhibitor, generated at 34 °C, is cooled for 10 min a either 25 °C or 0 °C, it is still just as active when assayed at 34 °C in the absence of hemin, but it is inactivated more rapidly by hemin at 34 °C. These findings provide additional support for the hypothesis that the hemin-reversible inhibitor is the physiological mediator by which hemin controls globin synthesis in intact reticulocytes and their cell-free systems.