Independent Signaling of grp78 Gene Transcription and Phosphorylation of Eukaryotic Initiation Factor 2α by the Stressed Endoplasmic Reticulum

Abstract

Perturbation of endoplasmic reticular (ER) function signals increased expression of the gene encoding the ER resident chaperone Grp78/BiP and rapid suppression of translational initiation accompanied by phosphorylation of the alpha-subunit of eukaryotic initiation factor 2 (eIF-2). eIF-2 alpha phosphorylation and grp78 mRNA induction were measured in GH3 pituitary cells subjected to varied degrees of ER stress to ascertain whether activation of an eIF-2 alpha kinase is involved in both events. grp78 mRNA was induced at low concentrations of ionomycin and dithiothreitol that did not provoke eIF-2 alpha phosphorylation or inhibition of amino acid incorporation. Mobilization of the bulk of cell-associated Ca2+ and the induction of grp78 mRNA occurred at comparable low concentrations of ionomycin, whereas phosphorylation of eIF-2 alpha and inhibition of protein synthesis required higher ionophore concentrations. Pretreatment for 1 h with cycloheximide suppressed grp78 mRNA induction and eIF-2 alpha phosphorylation in response to either stressor. Prolonged (17 h) cycloheximide blockade increased eIF-2 alpha phosphorylation without inducing grp78 mRNA. Upon release from the blockade, grp78 mRNA was induced and eIF-2 alpha was dephosphorylated. Translational tolerance to ionomycin or dithiothreitol, accompanied by dephosphorylation of eIF-2 alpha, was observed whenever grp78 mRNA was induced. Induction of grp78 mRNA preceded significant eIF-2 alpha phosphorylation during treatment with brefeldin A. It is concluded that signaling of grp78 gene transcription can occur independently of eIF-2 alpha phosphorylation or translational repression and that greater degrees of ER stress are required for eIF-2 alpha phosphorylation than for grp78 mRNA induction.

Immunoglobulin Binding Protein (BiP) Function Is Required to Protect Cells from Endoplasmic Reticulum Stress but Is Not Required for the Secretion of Selective Proteins

“…Cells respond to malfolded proteins by induction of GRP expression (2) and inhibition of protein synthesis (49). Upon calcium depletion from the ER, protein folding is disrupted and protein synthesis is inhibited by phosphorylation of the eukaryotic translation initiation factor eIF-2␣ (82).…”

“…Synthesis Mediated by A23187 Treatment-Agents that mediate ER stress, such as ionophore or tunicamycin treatment, elicit an immediate inhibition of protein synthesis initiation through phosphorylation of the eukaryotic translation initiation factor 2␣ subunit (44,49). We asked whether overexpression of BiP can influence the inhibition of protein synthesis in response to A23187 treatment.…”

Decreased ER-associated degradation of α-TCR induced by Grp78 depletion with the SubAB cytotoxin

“…Since phosphorylation of eIF2α is associated with a repression of protein synthesis (Brostrom et al, 1995), we decided to check how SubAB affects global rates of protein synthesis. Incorporation of 35 S-Met into newly synthesized proteins drops to ~5% of control levels 1 h after the addition of SubAB and returns to baseline levels 6 h after incubation with the toxin (Fig.…”

Regulation of Protein Synthesis in Ventricular Myocytes by Vasopressin

“…An adaptive response, characterized by translational accommodation to continued depletion of ER Ca 2ϩ stores by drugs such as ionomycin or thapsigargin or to the continued presence of a thiol-reducing agent, occurs over several hours. This adaptive response is dependent on increased expression of the ER resident chaperone, GRP78/BiP (1,4,7,8). Inductions of this chaperone and of recovery from translational inhibition depend on activation of grp78 transcription and, in some cell types, a growth-promoting factor.…”