In prostate cancer cells expression of C/EBPα down-regulates metallothionein and increases zinc toxicity Free

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The prostate harbors the highest zinc concentration of any tissue. In prostate cancer the zinc concentration decreases. Cultured prostate cancer cells undergo apoptosis in the presence of zinc. The mechanisms for the change in zinc metabolism in prostate cancer and the increased sensitivity of prostate cancer cells to zinc are unknown. The metallothioneins (MTs) are a family of zinc storage proteins which prevents cellular zinc toxicity. Regulation of expression of MT in prostate cells is poorly understood. MT levels decrease with prostate carcinogenesis and MT expression prevents cadmium induced prostate cancer. The CCAAT enhancer binding protein alpha (C/EBPα) is a transcription factor that plays an important role in regulating terminal differentiation of various cell types. C/EBPα is expressed in normal prostate epithelium but is absent in various prostate cancer cell lines. The role of C/EBPα in regulation of prostate development and carcinogenesis is unknown. Using transduction with a retrovirus we generated in three prostate cancer cell lines, PC3, LNCaP, and DU145, cell lines which overexpressed C/EBPα. We then examined the effect of forced expression of C/EBPα in these prostate cancer cells on global gene expression profile and found that C/EBPα significantly decreased RNA levels of MTs. Western blot analysis showed that C/EBPα reduced MT expression in both androgen-dependent and independent prostate cancer cells both under basal conditions and after zinc induction. MT is also induced by other metals including cadmium. The induction of MT by cadmium was not affected by C/EBPα expression in PC3 and LNCaP cells and only slightly reduced in DU145 cells. The MT promoter contains several C/EBPα consensus sequence binding sites suggesting that suppression of the expression of MTs may be from inhibition of the promoter activity of MTs by C/EBPα. Co-transfection of C/EBPα and the MT promoter construct in a luciferase reporter plasmid showed 1.5 to 2.5 fold inhibition of the MT promoter after exposure to zinc. Concomittant with the suppression of MT expression by C/EBPα the DU145 and LNCaP cells exhibited increasing sensitivity to zinc. For example after 3 days at 150 μM zinc, proliferation of LNCaP cells not expressing C/EBPα was 39.5 ± 5.5 % and decreased to 8.3 ± 3.2% in the C/EBPα expressing cells. Similarly, proliferation in the non-expressing DU145 cells decreased from 75.3 ± 6.3 % to 25.4 ± 13.1% in the C/EBPα expressing cells. Interestingly, PC3 cells overexpressing C/EBPα were more resistant to zinc with proliferation of 7.8 ± 3.1 % in the non-expressing cells increasing to 25.8 ± 5.2% in the C/EBPα expressing cells. Our data demonstrate that expression of MTs in prostate cancer cells is inhibited by C/EBPα and the effect may have functional significance in regulating the growth of prostate cancer cells and the response of these cells to environmental stress.