Calcium Ions and Phospholipid Activate a Translational Inhibitor in Reticulocyte Lysates

Abstract

Protein synthesis in rabbit reticulocyte lysates starts at a high rate but declines sharply within a few minutes unless the system is supplemented with hemin (London et al. 1976; Hunt et al. 1972; Mathews et al. 1973). Heme deficiency activates an inhibitor of protein synthesis initiation (heme controlled translational inhibitor or HCI) (Maxwell et al. 1971; Gross and Rabinovitz 1972), a cyclic AMP-independent protein kinase that specifically phosphorylates the small, α subunit of the initiation factor eIF-2 interfering with its function (Farrell et al. 1977; Levin et al. 1976; Kramer et al. 1976; Gross and Mendelewski 1977). There are a number of ways in which HCI can be activated (Ochoa 1983), including high hydrostatic pressure, elevated temperatures, sulfhydryl reagents (e.g., N-ethylmaleimide), or low levels of oxidized glutathione (GSSG), although the mechanism of activation is unknown in all cases. Only the activation due to heme deficiency or GSSG would appear to be physiologically relevant. Activation of HCI in lysates leads to phosphorylation of a polypeptide of M_r ~90 000 as well as of the eIF-2 α subunit. The 90 000 M_r polypeptide is HCI itself.