Tumor Suppressor Protein p53 mRNA and Subcellular Localization Are Altered by Changes in Cellular Copper in Human Hep G2 Cells

Abstract

Copper toxicity causes hepatic damage that can lead to the development of hepatocarcinoma. Similarly, copper deficiency has been reported to increase hepatocyte tumorigenesis. Thus, the objective of this work was to explore the role of copper toxicity and deficiency in the regulation of the tumor suppressor protein p53. Using Northern analysis, Western analysis, immunocytochemistry and the human hepatoma cell line Hep G2, this work showed that elevations in hepatocyte copper consistent with Wilson's disease (5.7-fold increase) induced p53 mRNA and confirmed that copper toxicity is correlated with apoptotic cell death. However, Western analysis and immunocytochemistry showed that post-transcriptional mechanisms are a significant part of the process, with p53 translocation from the cytosol into the nucleus of copper-treated cells. Treatment of Hep G2 cells with increasing concentrations of the copper chelator tetraethylenepentamine (TEPA, 0-50 micromol/L, 48 h) reduced cellular copper and increased mean p53 mRNA abundance by over fourfold with nuclear translocation of the wild-type protein. However, TEPA treatment did not result in a loss of cell viability or appear to induce apoptosis.

Transcriptional Activation of Stress Genes and Cytotoxicity in Human Liver Carcinoma (HepG2) Cells Exposed to Pentachlorophenol

“…The current study confirms previous works showing that various chemical compounds have the potential to up-regulate stress genes in human liver carcinoma (HepG 2 ) cells [26,29,30]. Exposure of HepG 2 cells to PCP revealed a significant induction of a number of stress genes including CYP1A1, XRE, HMTIIA, c-fos, and GADD153.…”

“…Exposure of HepG 2 cells to PCP revealed a significant induction of a number of stress genes including CYP1A1, XRE, HMTIIA, c-fos, and GADD153. Further, several studies report that metal and nonmetal compounds modulate transcriptional activation of stress genes in the human hepatoma cell line, HepG 2 [26,29,30]. Moreover, several reports in the literature have addressed the potential of organochlorine pesticides to initiate reporter gene transcription from estrogen response elements in DNA [32].…”

X-ray fluorescence microscopy reveals large-scale relocalization and extracellular translocation of cellular copper during angiogenesis

“…To resolve this issue, HMVEC tubulogenesis was initiated and the cells were exposed to three copper chelators: ammonium tetrathiomolybdate (TTM), tetraethylpentamine (TEPA), or bathocuproinedisulfonic acid (BCS). These chelators were chosen for their copper specificity and reported low toxicity in a number of cell types (10,26,27). Their low toxicity in HMVECs was confirmed by a trypan blue exclusion cell viability assay, which found no statistically significant decrease in viability from the use of any of the three chelators at the concentrations used in our experimental conditions.…”

Mechanisms of the Copper-dependent Turnover of the Copper Chaperone for Superoxide Dismutase

“…Although the qualitative changes in CCS abundance under the copper altered conditions used in this study were consistently reproduced, differences in the absolute amount of CCS were observed (see Table 1). Because the cell number and media requirements were kept identical in each experiment, this variation in the absolute amount of CCS under such conditions likely reflects reported differences in the amounts of available copper for chelation in unique subcellular locations that may change during physiological events such as the cell cycle (31). Consistent with this concept, statistical analysis of the -fold differences in Table 1 illustrates that C244S,C246S CCS was the only mutation where the change in CCS abundance with copper content was statistically significant when compared with the endogenous CCS ϩ/ϩ cells.…”