Thapsigargin-induced persistent intracellular calcium pool depletion and apoptosis in human hepatoma cells

Kaneko, Yoshiyasu; Tsukamoto, Ayumi

doi:10.1016/0304-3835(94)90253-4

Cited by 85 publications

(45 citation statements)

References 17 publications

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“…In addition to tunicamycin, ER stress was induced by the sarcoplasmic-endoplasmic reticulum calcium ATPase inhibitor thapsigargin, which depletes intraluminal ER calcium and induces apoptosis (42). As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Endoplasmic Reticulum Stress-induced Cysteine Protease Activation in Cortical Neurons

Siman

Flood²,

Wang

et al. 2001

Journal of Biological Chemistry

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Endoplasmic reticulum (ER) stress elicits protective responses of chaperone induction and translational suppression and, when unimpeded, leads to caspase-mediated apoptosis. Alzheimer's disease-linked mutations in presenilin-1 (PS-1) reportedly impair ER stress-mediated protective responses and enhance vulnerability to degeneration. We used cleavage site-specific antibodies to characterize the cysteine protease activation responses of primary mouse cortical neurons to ER stress and evaluate the influence of a PS-1 knock-in mutation on these and other stress responses. Two different ER stressors lead to processing of the ER-resident protease procaspase-12, activation of calpain, caspase-3, and caspase-6, and degradation of ER and non-ER protein substrates. Immunocytochemical localization of activated caspase-3 and a cleaved substrate of caspase-6 confirms that caspase activation extends into the cytosol and nucleus. ER stress-induced proteolysis is unchanged in cortical neurons derived from the PS-1 P264L knock-in mouse. Furthermore, the PS-1 genotype does not influence stress-induced increases in chaperones Grp78/BiP and Grp94 or apoptotic neurodegeneration. A similar lack of effect of the PS-1 P264L mutation on the activation of caspases and induction of chaperones is observed in fibroblasts. Finally, the PS-1 knock-in mutation does not alter activation of the protein kinase PKR-like ER kinase (PERK), a trigger for stress-induced translational suppression. These data demonstrate that ER stress in cortical neurons leads to activation of several cysteine proteases within diverse neuronal compartments and indicate that Alzheimer's disease-linked PS-1 mutations do not invariably alter the proteolytic, chaperone induction, translational suppression, and apoptotic responses to ER stress.

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Section: Resultsmentioning

confidence: 99%

Endoplasmic Reticulum Stress-induced Cysteine Protease Activation in Cortical Neurons

Siman

Flood²,

Wang

et al. 2001

Journal of Biological Chemistry

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“…The specific mechanism whereby mutant PS-1 disrupts cellular calcium homeostasis is unknown but may involve an effect on ER calcium regulation, as suggested by the localization of PS-1 in the ER (14) and the demonstrations of enhanced calcium responses to agonists that induce calcium release from ER in PC12 cells expressing mutant PS-1 (45) and in fibroblasts from human carriers of PS-1 mutations (55). Indeed, calcium release from ER induced by the ER Ca 2ϩ -ATPase inhibitor thapsigargin is sufficient to trigger apoptosis in several types of cells (56,57).…”

Section: Discussionmentioning

confidence: 99%

Calbindin D28k blocks the proapoptotic actions of mutant presenilin 1: Reduced oxidative stress and preserved mitochondrial function

Guo

Christakos

Robinson

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

208

179

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Mutations in the presenilin 1 (PS-1) gene account for many cases of early-onset autosomal dominant inherited forms of Alzheimer's disease. Recent findings suggest that PS-1 mutations may sensitize neurons to apoptosis induced by trophic factor withdrawal and exposure to amyloid ␤-peptide (A␤). We now report that overexpression of the calcium-binding protein calbindin D28k prevents apoptosis in cultured neural cells expressing mutant PS-1 (L286V and M146V missense mutations). Elevations of the intracellular Ca 2؉ concentration and generation of reactive oxygen species induced by A␤, and potentiated by mutant PS-1, were suppressed in calbindin-overexpressing cells. Impairment of mitochondrial function by A␤ (which preceded apoptosis) was exacerbated by PS-1 mutations and was largely prevented by calbindin. These findings suggest that PS-1 mutations render neurons vulnerable to apoptosis by a mechanism involving destabilization of cellular calcium homeostasis, which leads to oxidative stress and mitochondrial dysfunction.Degeneration and death of neurons in brain regions involved in learning and memory processes underlie Alzheimer's disease (AD), a fatal disorder that affects millions of persons worldwide. Although the cause(s) of the vast majority of cases of AD are unknown, a small percentage of cases are inherited in an autosomal dominant manner. Missense mutations in three different genes, ␤-amyloid precursor protein (APP; chromosome 21), presenilin 1 (PS-1; chromosome 14), and PS-2 (chromosome 1) have been causally linked to early-onset familial forms of AD (1). APP is a transmembrane protein that is the source of the 40-to 42-aa amyloid ␤-peptide (A␤) that forms fibrillar aggregates (plaques) associated with degenerating neurons in the brain in AD. Several lines of evidence suggest a major role for A␤ in the neurodegenerative process in AD as follows: APP mutations result in increased production of cytotoxic forms of A␤ (2); transgenic mice expressing mutated human APP exhibit A␤ deposition and cognitive impairments (3, 4); and A␤ damages and kills cultured neurons by a mechanism involving oxidative stress and disruption of cellular calcium homeostasis (5-10). PS-1 and PS-2 encode integral membrane proteins with six or eight membrane-spanning domains (11-13) and are localized in the endoplasmic reticulum (ER; ref. 14). PSs are expressed in neurons throughout the brain (15-17) and appear to be present in both degenerating and nondegenerating neurons in AD brain (18,19). Mutations in PS-1 account for approximately 50% of all cases of familial AD (20); understanding how PS-1 mutations promote neuron degeneration is, therefore, a critical issue in AD research.Increased oxidative stress and disruption of neuronal calcium homeostasis appear to be interrelated final common pathways that mediate the neurodegenerative process in AD (for review, see refs. 21 and 22). There is increased protein, lipid, and DNA oxidation in association with degenerating neurons in AD (23-27). Exposure of cultured neurons to A␤ in...

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“…Ca 2ϩ ionophores or the sarcoplasmic reticulum Ca 2ϩ -ATPase pump inhibitor, thapsigargin, TG) can trigger apoptosis in diverse cell types (23)(24)(25)(26)(27). Inhibition of the sarcoplasmic reticulum Ca 2ϩ -ATPase pump by TG causes a transient increase in cytoplasmic Ca 2ϩ from ER Ca 2ϩ stores, and a later influx of Ca 2ϩ from the extracellular milieu, leading to the induction of apoptotic cell death (24,27,28). Consequently, emptying of intracellular Ca 2ϩ stores may trigger apoptosis by disrupting the intracellular architecture and allowing key elements of the effector machinery (e.g.…”

Section: ␤-Lapmentioning

confidence: 99%

Calcium Is a Key Signaling Molecule in β-Lapachone-mediated Cell Death

Tagliarino¹,

Pink²,

Dubyak

et al. 2001

Journal of Biological Chemistry

153

150

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␤-Lapachone (␤-Lap) triggers apoptosis in a number of human breast and prostate cancer cell lines through a unique apoptotic pathway that is dependent upon NQO1, a two-electron reductase. Downstream signaling pathway(s) that initiate apoptosis following treatment with ␤-Lap have not been elucidated. Since calpain activation was suspected in ␤-Lap-mediated apoptosis, we examined alterations in Ca 2؉ homeostasis using NQO1-expressing MCF-7 cells. ␤-Lap-exposed MCF-7 cells exhibited an early increase in intracellular cytosolic Ca 2؉, from endoplasmic reticulum Ca 2؉ stores, comparable to thapsigargin exposures. 1,2-Bis-(2-aminophenoxy)ethane-N,N,N,N-tetraacetic acid-acetoxymethyl ester, an intracellular Ca 2؉ chelator, blocked early increases in Ca 2؉ levels and inhibited ␤-Lap-mediated mitochondrial membrane depolarization, intracellular ATP depletion, specific and unique substrate proteolysis, and apoptosis. The extracellular Ca 2؉ chelator, EGTA, inhibited later apoptotic end points (observed >8 h, e.g. substrate proteolysis and DNA fragmentation), suggesting that later execution events were triggered by Ca 2؉ influxes from the extracellular milieu. Collectively, these data suggest a critical, but not sole, role for Ca 2؉ in the NQO1-dependent cell death pathway initiated by ␤-Lap. Use of ␤-Lap to trigger an apparently novel, calpain-like-mediated apoptotic cell death could be useful for breast and prostate cancer therapy. ␤-Lap1 is a naturally occurring compound present in the bark of the South American Lapacho tree. It has antitumor activity against a variety of human cancers, including colon, prostrate, promyelocytic leukemia, and breast (1-3). ␤-Lap was an effective agent (alone and in combination with taxol) against human ovarian and prostate xenografts in mice, with little host toxicity (4). We recently demonstrated that ␤-Lap kills human breast and prostate cancer cells by apoptosis, a cytotoxic response significantly enhanced by NAD(P)H:quinone oxidoreductase (NQO1, E.C. 1.6.99.2) enzymatic activity (5).2 ␤-Lap cytotoxicity was prevented by co-treatment with dicumarol (an NQO1 inhibitor) in NQO1-expressing breast and prostate cancer cells (5).2 NQO1 is a cytosolic enzyme elevated in breast cancers (6) that catalyzes a two-electron reduction of quinones (e.g. ␤-Lap, menadione), utilizing either NADH or NADPH as electron donors. Reduction of ␤-Lap by NQO1 presumably leads to a futile cycling of the compound, wherein the quinone and hydroquinone form a redox cycle with a net concomitant loss of reduced NAD(P)H (5).Apoptosis is an evolutionarily conserved pathway of biochemical and molecular events that underlie cell death processes involving the stimulation of intracellular zymogens. The process is a genetically programmed form of cell death involved in development, normal turnover of cells, and in cytotoxic responses to cellular insults. Once apoptosis is initiated, biochemical and morphological changes occur in the cell. These changes include: DNA fragmentation, chromatin condensation, cytoplasmic membran...

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Thapsigargin-induced persistent intracellular calcium pool depletion and apoptosis in human hepatoma cells

Cited by 85 publications

References 17 publications

Endoplasmic Reticulum Stress-induced Cysteine Protease Activation in Cortical Neurons

Endoplasmic Reticulum Stress-induced Cysteine Protease Activation in Cortical Neurons

Calbindin D28k blocks the proapoptotic actions of mutant presenilin 1: Reduced oxidative stress and preserved mitochondrial function

Calcium Is a Key Signaling Molecule in β-Lapachone-mediated Cell Death

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