Endoplasmic reticulum stress proteins block oxidant-induced Ca2+ increases and cell death.
Oxidants are important human toxicants. Increased intracellular free Ca2+ may be critical for oxidant toxicity, but this mechanism remains controversial. Furthermore, oxidants damage the endoplasmic reticulum (ER) and release ER Ca2+, but the role of the ER in oxidant toxicity and Ca2+ regulation during toxicity is also unclear. tert-Butylhydroperoxide (TBHP), a prototypical organic oxidant, causes oxidative stress and an increase in intracellular free Ca2+. Therefore, we addressed the mechanism of oxidant-induced cell death and investigated the role of ER stress proteins in Ca2+ regulation and cytoprotection after treating renal epithelial cells with TBHP. Prior ER stress induces expression of the ER stress proteins Grp78, Grp94, and calreticulin and rendered cells resistant to cell death caused by a subsequent TBHP challenge. Expressing antisense RNA targeted to grp78 prevents grp78 induction sensitized cells to TBHP and disrupted their ability to develop cellular tolerance. In addition, overexpressing calreticulin, another ER chaperone and Ca2+-binding protein, also protected cells against TBHP. Interestingly, neither prior ER stress nor calreticulin expression prevented lipid peroxidation, but both blocked the rise in intracellular free Ca2+ after TBHP treatment. Loading cells with EGTA, even after peroxidation had already occurred, also prevented TBHP-induced cell death, indicating that buffering intracellular Ca2+ prevents cell killing. Thus, Ca2+ plays an important role in TBHP-induced cell death in these cells, and the ER is an important regulator of cellular Ca2+ homeostasis during oxidative stress. Given the importance of oxidants in human disease, it would appear that the role of ER stress proteins in protection from oxidant damage warrants further consideration.
Liu
H
H
Adirondack Biomedical Research Institute, Lake Placid, New York 12946, USA.
Miller
E
E
van de Water
B
B
Stevens
J L
JL
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D037282:Calreticulin; D002352:Carrier Proteins / Q000378:metabolism*; D016923:Cell Death; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006360:Heat-Shock Proteins; D018374:LLC-PK1 Cells; D008565:Membrane Proteins / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology*; D010545:Peroxides / Q000494:pharmacology; D012261:Ribonucleoproteins / Q000378:metabolism*; D013552:Swine; D020122:tert-Butylhydroperoxide
D002135:Calcium-Binding Proteins; D037282:Calreticulin; D002352:Carrier Proteins; D000091342:Endoplasmic Reticulum Chaperone BiP; D018840:HSP70 Heat-Shock Proteins; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D008565:Membrane Proteins; D018832:Molecular Chaperones; D016877:Oxidants; D010545:Peroxides; D012261:Ribonucleoproteins; C018551:glucose-regulated proteins; D020122:tert-Butylhydroperoxide; D002118:Calcium