Translational Control of Protein Synthesis in Reticulocyte Lysates by eIF-2α Kinases

Abstract

The initiation of protein synthesis in reticulocyte lysates is inhibited by heme deficiency, double-stranded RNA, or oxidized glutathione (Zucker and Schulman 1968; Rabinovitz et al. 1969; Hunt et al. 1972; Ehrenfeld and Hunt 1971; Kosower et al. 1972). In lysates, the inhibitions are all characterized by a brief period of control linear synthesis, followed by an abrupt decline in this rate as protein synthesis shuts off. The inhibitions are due primarily to the activation of cAMP-independent protein kinases that specifically phosphorylate the 38000 mol. wt. α-subunit of the eukaryotic initiation factor-2 (eIF-2) (Levin et al. 1976; Kramer et al. 1976;Ranu and London 1976; Farrell et al. 1977; Gross and Mendelewski 1977; Levin and London 1978; Ernst et al. 1978a). Similar inhibitions of protein synthesis are produced by adding the purified inhibitors, the heme-regulated eIF-2α kinase (HRI), or the double-stranded RNA-activated eIF-2α kinase (dsI), to reticulocyte lysates. The inhibition of protein synthesis is a result of a block in the early steps of protein chain initiation and is immediately preceded by a decrease in the formation of the [eIF-2 Met-tRNA_f·GTP] ternary complex and the [40S-eIF-2-Met-tRNA_f·GTP] 43S initiation complex (de Haro et al. 1978; Ranu et al. 1978; de Haro and Ochoa 1978; Ranu and London 1979; de Haro and Ochoa 1979; Das et al. 1979). Studies by Cherbas and London (1976) suggested that the principal effect of HRI was the impairment of the recycling of eIF-2. In vitro studies have suggested that the recycling of eIF-2 requires the presence of an additional multipolypeptide factor, whose function is inhibited by the phosphorylation of eIF-2.