Enhanced Apoptosis in Metallothionein Null Cells
- Yukihiro Kondo1,
- James M. Rusnak,
- Dale G. Hoyt,
- Catherine E. Settineri,
- Bruce R. Pitt and
- John S. Lazo
- Department of Pharmacology, University of Pittsburgh and Experimental Therapeutics Program, University of Pittsburgh Cancer Institute, Pittsburgh, Pennsylvania 15261
Abstract
Metallothioneins (MTs) are major intracellular, zinc-binding proteins with antioxidant properties. Mouse embryonic cells null for MT due to loss of functional MT I and II genes (MT−/−) were more susceptible to apoptotic death after exposure to tert-butyl hydroperoxide or the anti-cancer agents cytosine arabinoside, bleomycin, melphalan, andcis-dichlorodiammineplatinum(II) compared with wild-type mouse embryonic cells (MT+/+). We measured basal levels of the tumor suppressor protein p53 and the death effector protein Bax and found the basal levels of both proteins were higher in MT null cells compared with MT+/+ cells. After treatment with the DNA-damaging agentcis-dichlorodiammineplatinum(II), p53 protein levels were induced in both MT+/+ and MT−/− cells with MT null cells always maintaining the highest p53 levels. The elevated sensitivity to apoptosis was not restricted to embryonic cells. Primary pulmonary fibroblasts were isolated from distinct litters of MT null, heterozygous, and wild-type mice, and all had undetectable basal MT levels. Zinc exposure increased MT levels in the wild-type and heterozygous fibroblasts but not in the MT null fibroblasts. Consistent with the induced MT levels, we found MT+/+ and MT+/− embryonic cells were less sensitive tocis-dichlorodiammineplatinum(II)-induced apoptosis compared with MT−/− cells. Our results implicate MT as a stress-responsive factor that can regulate apoptotic engagement.
Footnotes
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Send reprint requests to: Prof. John S. Lazo, Chairman, Department of Pharmacology, University of Pittsburgh School of Medicine, E1340 Biomedical Science Tower, Pittsburgh, PA 15261.
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↵1 Current affiliation: Department of Urology, Nippon Medical School, Bunkyo-ku, Tokyo, Japan, 113.
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This research was funded in part by American Institute for Cancer Research Grant 95A50), the American Heart Association, American Cancer Society Grant IRG-58–34 and National Institutes of Health Grants CA61299, DK46935, and HL32154.
- Abbreviations:
- MT
- metallothionein
- ROI
- reactive oxygen intermediates, ara-C, cytosine arabinoside
- tBH
- tert-butyl hydroperoxide
- CDDP
- cis-dichlorodiammineplatinum(II) (cisplatin)
- QFIGE
- quantitative field inversion gel electrophoresis
- PBS
- phosphate-buffered saline
- MEC
- mouse embryonic cells
- BLM
- bleomycin
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- Received October 29, 1996.
- Accepted May 2, 1997.
- The American Society for Pharmacology and Experimental Therapeutics
