Involvement of PKCδ, p38, p53 and Gadd45 Signaling Pathway Delayed G2/M Cell Cycle Progression in Zinc Supplemented Normal Human Bronchial Epithelial Cells

Abstract

The involvement of PKCδ, p38, p53 and Gadd45 signal pathway on G2/M cell cycle progression was examined in normal human bronchial cells (NHBE) cultured in ZD (zinc deficient, <1μM), ZN (zinc normal, 4μM), ZA (zinc adequate, 16μM) and ZS (zinc supplementation, 32μM) media. A blockage of G2/M cell cycle progression was accompanied by markedly increases in Gadd45 and p53 mRNA and protein levels in ZS than ZN control cells. siRNA-mediated knocking down of Gadd45 relieved G2/M blockage and triggered apoptosis, suggesting that Gadd45 is responsible for the G2/M cell cycle blockage and protection of cells from apoptosis, which is caused by possible genotoxic stress associated with high cellular zinc status. Interestingly, p53 protein expression was depressed after knocked down Gadd45. Administration of p53 inhibitor, pifithrin, also relieved the blockage in G2/M, suggesting that p53 and Gadd45 may form a positive feedback loop in the delay of G2/M in ZS cells. Moreover, twofold higher p38 mRNA and PKCδ protein levels were observed in ZS than ZN cells. Furthermore, the inhibition of p38 and PKCδ by specific protein inhibitors, SB202190 and Rottlerin, respectively, as well as by the overexpression of p38 and PKCδ dominant-negatives, prevented the zinc supplementation induced G2/M blockage in NHBE cells. Thus, in ZS NHBE cells, the upregulation of Gadd45, may be modulated by enhanced PKCδ, p38 and p53 expression, which led to the delay in G2/M cell cycle progression.