Submitted to: Experimental Biology and Medicine
Publication Type: Peer reviewed journal
Publication Acceptance Date: 1/22/2008
Publication Date: 3/2/2008
Citation: Shih, R.S., Wong, S.H., Schoene, N.W., Lei, K.Y. 2008. Suppression of Gadd45 Relieves the G2/M Cell Cycle Blockage and Enhanced Phosphorylation of p53 and p38 in Zinc Supplemented Normal Human Bronchial Epithelial Cells. Experimental Biology and Medicine. 233:317-327. Interpretive Summary: Zinc is an essential nutrient necessary for the synthesis and function of many proteins critical for the regulation of proper cellular responses to stresses. However, exposure to high amounts of zinc can be detrimental, especially to lung cells, producing imbalances in actions of stress proteins which may favor inflammatory and/or pre-cancerous conditions. This experiment with cultured normal cells from the lung focused on studying the effects of different amounts of zinc on the expression of a protein (Gadd45) that plays an critical role in sensing the amount of stress in the cell. Cells were grown in media containing 0.4, 4, 16, and 32 micromoles of zinc. With the highest amount of zinc, both messenger RNA and protein for Gadd45 were significantly increased compared to the values observed in cells grown in 0.4, 4, and 16 micromoles of zinc. Two other stress-detecting proteins also demonstrated enhanced phosphorylation in cells grown in the highest amount of zinc. The results provide evidence that high amounts of zinc in cell media can create imbalances in cellular stress responses that produce adverse effects in normal cells. These are novel observations and provide information to nutritionists and other researchers about potential problems that may be produced by zinc excess.
Technical Abstract: An adequate zinc status is essential for optimal cellular functions and growth. Yet, excessive zinc supplementation can be cytotoxic and can impair cell growth. Gadd45 plays a vital role as a cellular stress sensor in the modulation of cell signal transduction in response to stress. The present study was designed to determine the influence of zinc status on Gadd45 expression and cell cycle progression in zinc deficient and supplemented normal human bronchial epithelial (NHBE) cells, and to decipher the molecular mechanism(s) exerted by the suppression of Gadd45 expression on cell growth and cell cycle progression in this cell type. Cells were cultured for one passage in different concentrations of zinc: <0.4 microM (ZD) as severe zinc deficient; 4 microM as a normal zinc level in culture media; 16 microM (ZA) as the normal human plasma zinc level; and 32 microM (ZS) as the high end of plasma zinc possibly attainable by extreme oral supplementation. Inhibition of cell growth, up-regulation of Gadd45 mRNA and protein expression, and blockage of G2/M cell cycle progression were observed in ZS cells. In contrast, little or no changes in these parameters were seen in ZD cells. The siRNA-mediated knocking down of Gadd45 was found to relieve G2/M blockage in ZS cells, which indicated that the blockage was Gadd45 dependent. Moreover, the enhanced phosphorylation of p38 and p53 (ser15) in ZS cells was normalized after suppression of Gadd45 by siRNA, indicating that the enhanced phosphorylation of these proteins was Gadd45 dependent. Thus, we demonstrated for the first time that elevated zinc levels modulated signal transduction to produce a delay at G2/M during cell cycle progression in NHBE cells.