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Resistance to tumor necrosis factor-induced cell death mediated by PMCA4 deficiency
1Department of Immunology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Abstract:
We used retrovirus insertion-mediated random mutagenesis to generate tumor necrosis factor (TNF)-resistant lines from L929 cells. Using this approach, we discovered that the plasma membrane calcium ATPase 4 (PMCA4) is required for TNF-induced cell death in L929 cells. Under basal conditions, PMCA4-deficient (PMCA(mut)) cells have a normal phenotype. However, stimulation with TNF induces an abnormal increase in the intracellular calcium concentration ([Ca(2+)](i)). The substantially elevated [Ca(2+)](i) caused resistance to TNF-induced cell death. We found that an increase in the total volume of acidic compartments (VAC), mainly constituted by lysosomes, is a common event in cell death caused by a variety of agonists. The increased [Ca(2+)](i) in PMCA(mut) cells promoted lysosome exocytosis, which, at least in part, accounted for the inhibition of TNF-induced increase in VAC and cell death. Promoting lysosome exocytosis by calcium inhibited TNF-induced cell death in wild-type L929 cells, while inhibition of lysosome exocytosis or increase of VAC by sucrose restored the sensitivity of PMCA(mut) cells to TNF-induced cell death. Thus, increase of the volume of acidic compartment is a part of the cell death process, and the antideath effect of calcium is mediated, at least in part, by inhibition of the TNF-induced increase in VAC.
Insights
Plasma membrane calcium ATPase 4 (PMCA4) deficiency causes resistance to tumor necrosis factor (TNF)-induced cell death by altering intracellular calcium levels. This resistance involves changes in lysosome function and acidic compartment volume.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Tumor necrosis factor (TNF) is a key mediator of cellular apoptosis and inflammation.
- Intracellular calcium homeostasis plays a critical role in regulating various cellular processes, including cell death.
- Lysosomes and acidic compartments are involved in cellular degradation and signaling pathways.
Purpose of the Study:
- To identify genes involved in resistance to TNF-induced cell death in L929 cells.
- To elucidate the role of plasma membrane calcium ATPase 4 (PMCA4) in TNF-induced apoptosis.
- To investigate the relationship between intracellular calcium, lysosome function, and cell death.
Main Methods:
- Retrovirus insertion-mediated random mutagenesis to generate TNF-resistant L929 cell lines.
- Phenotypic analysis of PMCA4-deficient cells under basal and TNF-stimulated conditions.
- Measurement of intracellular calcium concentration ([Ca(2+)](i)) and acidic compartment volume (VAC).
- Assessment of lysosome exocytosis and its impact on cell death.
Main Results:
- PMCA4-deficient cells exhibit resistance to TNF-induced cell death.
- TNF stimulation leads to an abnormal increase in intracellular calcium ([Ca(2+)](i)) in PMCA4-deficient cells.
- Increased [Ca(2+)](i) promotes lysosome exocytosis, inhibiting TNF-induced increase in acidic compartment volume (VAC) and cell death.
- Calcium-mediated lysosome exocytosis inhibits TNF-induced cell death in wild-type cells.
Conclusions:
- Plasma membrane calcium ATPase 4 (PMCA4) is essential for TNF-induced cell death in L929 cells.
- Elevated intracellular calcium in PMCA4-deficient cells confers resistance to TNF by modulating lysosome exocytosis and acidic compartment volume.
- Modulation of lysosome exocytosis and acidic compartment volume represents a novel mechanism influencing TNF-mediated cell death.