| Acquisition of Apoptotic Resistance in Cadmium-Transformed Human Prostate Epithelial Cells: Bcl-2 Overexpression Blocks the Activation of JNK Signal Transduction Pathway Wei Qu,1 Hengning Ke,2 Jingbo Pi,1 Daniel Broderick,1 John E. French,2 Mukta M. Webber,3 and Michael P. Waalkes1 1Inorganic Carcinogenesis Section, Laboratory of Comparative Carcinogenesis, National Cancer Institute at the National Institute of Environmental Health Sciences, and 2Laboratory of Molecular Toxicology, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina, USA; 3Departments of Zoology and Medicine, Michigan State University, East Lansing, Michigan, USA Abstract
Background: We have recently shown that cadmium can induce malignant transformation of the human prostate epithelial cell line (RWPE-1) and that these cadmium-transformed prostate epithelial (CTPE) cells acquire apoptotic resistance concurrently with malignant phenotype. Objective: The present study was designed to define the mechanism of acquired apoptotic resistance in CTPE cells. Methods: Various molecular events associated with apoptosis were assessed in control and CTPE cells that were obtained after 8 weeks of continuous cadmium exposure. Results: Compared with control, CTPE cells showed a generalized resistance to apoptosis induced by cadmium, cisplatin, or etoposide. Signal-regulated mitogen-activated protein kinases, extracellular signal-regulated kinases 1 and 2, c-Jun N-terminal kinases (JNK1 and JNK2) , and p38 were phosphorylated in a cadmium concentration-dependent fashion in CTPE and control cells. However, phosphorylated JNK1/2 levels and JNK kinase activity were much lower in CTPE cells. The pro-apoptotic gene Bax showed lower transcript and protein levels, whereas the anti-apoptotic gene Bcl-2 showed higher levels in CTPE cells. The ratio of Bcl-2/Bax, a key determinant in apoptotic commitment, increased more than 4-fold in CTPE cells. In Bcl-2–transfected PT-67 cells, phosphorylated JNK1/2 levels were much lower after apoptogenic stimulus, and apoptosis induced by cadmium or etoposide was reduced compared with control. Mutation of tyrosine to serine at the 21st amino acid of the Bcl-2 protein BH4 domain resulted in a loss both of suppression of JNK1/2 phosphorylation and its anti-apoptotic function. Conclusions: CTPE cells become resistant to apoptosis during malignant transformation, and disruption of the JNK pathway and Bcl-2 overexpression play important roles in this resistance. Bcl-2 BH4 domain is required for modulating JNK phosphorylation and anti-apoptotic function. Key words: apoptosis, Bcl-2, cadmium, JNK, malignant transformation. Environ Health Perspect 115:1094–1100 (2007) . doi:10.1289/ehp.10075 available via http://dx.doi.org/ [Online 5 April 2007]
Address correspondence to M.P. Waalkes, Inorganic Carcinogenesis Section, NCI at NIEHS, PO Box 12233, Mail Drop F0-09, 111 Alexander Dr., Research Triangle Park, NC 27709 USA. Telephone: (919) 541-2328. Fax: (919) 541-3970. E-mail: waalkes@niehs.nih.gov We thank L.K. Keefer, J. Liu, and E. Tokar for critical evaluation of this manuscript. This research was supported (in part) by the Intramural Research Program of the Center for Cancer Research, NCI, NIH. The authors declare they have no competing financial interests. Received 12 January 2007 ; accepted 5 April 2007.
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