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

Abstract

BiP/GRP78 is a lumenal stress protein of the endoplasmic reticulum (ER) that interacts with polypeptide folding intermediates transiting the secretory compartment. We have studied the secretion and the stress response in Chinese hamster ovary (CHO) cells that overexpress either wild-type immunoglobulin binding protein (BiP) or a BiP deletion molecule (residues 175-201) that can bind peptides and ATP but is defective in ATP hydrolysis and concomitant peptide release. Overexpressed wild-type BiP was localized to the ER and unique vesicles within the nucleus, whereas overexpressed ATPase-defective BiP was localized to the ER and cytoplasmic vesicles but was absent from the nucleus. Compared with wild-type CHO cells, overexpression of ATPase-defective BiP prevented secretion of factor VIII, a coagulation factor that extensively binds BiP in the lumen of the ER. Under these conditions factor VIII was stably associated with the ATPase-defective BiP. In contrast, the secretion of monocyte/macrophage colony stimulating factor, a protein that is not detected in association with BiP, was not affected by overexpression of ATPase-defective BiP. These results show that BiP function is not required for secretion of some proteins and suggest that some proteins do not interact with BiP upon transport through the ER.The The endoplasmic reticulum is the site where folding occurs for newly synthesized proteins that are destined for the cell surface. In addition, it is the principal cellular storage site for calcium. Calcium homeostasis is required for proper polypeptide folding and secretion of selective proteins (1) as well as for intracellular signaling events that occur within the cell. Within the ER 1 are resident cellular proteins, such as the glucoseregulated protein of 78 kDa (2), that are also known as the immunoglobulin-binding protein (BiP) (3, 4), the glucose-regulated protein of 94 kDa (GRP94), calnexin, calreticulin, and ERp72 that associate with folding intermediates. It is proposed that these cellular ER proteins act as chaperones to prevent aggregation of polypeptide folding intermediates (5). In addition, several of these proteins are documented to be the major calcium-binding proteins in the cell (1, 6, 7). Thus, the processes that maintain calcium homeostasis and proper polypeptide folding are likely to be intimately intertwined.Like all hsp70 family members, BiP binds ATP tightly (8) and has a weak ATPase activity (9) that is stimulated in vitro by small hydrophobic peptides (10) and that induces the release of BiP from bound polypeptides (4). Depletion of cellular ATP inhibits protein folding (11) and results in prolonged association of some proteins with BiP (12), whereas lowering ER calcium levels causes the release of T cell receptor subunits from BiP in vivo, presumably by inappropriately activating ATP hydrolysis (13). These results, together with the recent demonstration of ATP in the ER (14) and an ER ATP transport system (15), have provided evidence that the ATPase activity of BiP is important for its ...

Homocysteine-induced endoplasmic reticulum stress causes dysregulation of the cholesterol and triglyceride biosynthetic pathways

“…It follows that by blocking or minimizing ER stress, it may be possible to attenuate the induction of lipid biosynthesis. In support of this concept, we observed that stable overexpression of GRP78/BiP, which protects cells from agents or conditions known to cause ER stress (39,40), inhibits homocysteine-induced expression of genes involved in cholesterol biosynthesis in cultured human cells.…”

“…Overexpression of GRP78/BiP has been reported to protect cells from ER stress (39,40). Stably transfected cells overexpressing GRP78/BiP were generated to assess whether blocking homocysteine-induced ER stress attenuated the expression of SREBP-1 and IPP isomerase.…”

Thyroglobulin Transport along the Secretory Pathway

“…Moreover, in cells with increased BiP levels, diminished secretion of factor VIII or von Willebrand factor has been observed (39,60). Importantly, selectively increased ER chaperone expression does not produce a generalized effect on anterograde protein traffic, as export is slowed only for those secretory proteins to which chaperone binding can be detected (39,61). In the present study, Tg, a protein shown to bind GRP94, exhibits similarly delayed export from cells with increased levels of GRP94.…”

Preconditioning with Endoplasmic Reticulum Stress Ameliorates Mesangioproliferative Glomerulonephritis

“…2,3,5,26 Cells overexpressing GRP78 are resistant to conditions associated with ER stress. 27 The subnephritogenic dosages of tunicamycin and thapsigargin used for preconditioning in this study evoked the adaptive UPR without renal pathologic or functional effects in healthy control rats and ameliorated the severity of the disease manifestations of anti-Thy1 nephritis.…”