Abstract
Lead (Pb) and mercury (Hg) are widespread environmental contaminants that induce prominent neural toxicity. Although the brain is not the major Pb and Hg depot in the body, these metals preferentially accumulate in astroglia to exert toxic effects. In this study, we examined the effects of Pb acetate and HgCl2 on the expression of GRP78, a molecular chaperone in the endoplasmic reticulum (ER) that may provide cytoprotection in response to cellular stresses in the C6 rat glioma cell line. We also evaluated the DNA binding activities of several redox-regulated transcription factors in metal-treated cells. Our results showed that mRNA levels of GRP78 were up-regulated by Pb and Hg at 0.1 and 1 μM, but down-regulated at higher concentrations (10 μM). GRP78 protein levels increased in a concentration- and time-dependent manner in Pb and/or Hg-treated cells. Pb increased protein binding to the GST-Ya antioxidant/electrophile response element (ARE/EpRE) and to the NF-kB consensus binding sequence of the cytomegalovirus 2 (CMV2) promoter, but decreased protein binding to the Ha-ras ARE/EpRE or to the c-fos 12-O-tetradecanoyl-phorbol-13-acetate (TPA) response element (TRE). In contrast, Hg activated DNA binding by all redox-regulated transcription factors. These studies shed some light on the molecular mechanisms of Pb and Hg toxicity in C6 rat glioma cells and suggest that GRP78 and oxidative stress may participate in the neurotoxic response to these metals.
Similar content being viewed by others
References
Aschner, M., Rising, L. and Mullaney, K. (1996) “Differential sensitivity of neonatal rat astrocyte cultures to mercuric chloride (MC) and methylmercury (MeHg): Studies on K+ and amino acid transport and metallothionein (MT) induction”,Neurotoxicology 17, 107–116.
Bertolotti, A., Zhang, Y., Hendershot, L.M., Harding, H.P. and Ron, D. (2000) “Dynamic interaction of BiP and ER stress transducers in the unfolded-protein response”,Nature Cell. Biol. 2, 326–332.
Bissell, M.G., Eng, L.F., Herman, M.M., Bensch, K.G. and Miles, L.E. (1975) “Quantitative increase of neuroglia-specific GFA protein in rat C-6 glioma cells in vitro”,Nature 255, 633–634.
Bral, CM. and Ramos, K.S. (1997) “Identification of benzo[a]pyrene-induciblecu-acting elements within c-Ha-ras transcriptional regulatory sequences”,Mol. Pharmacol. 52, 974–982.
Charleston, J.S., Body, R.L., Mottet, N.K., Vahter, M.E. and Burbacher, T.M. (1995) “Autometallographic determination of inorganic mercury distribution in the cortex of the calcarrne sulcus of the monkeyMacaca fascicularis following long-term subclinical exposure to methylmercury and mercuric chloride”,Toxicol. Appl. Pharmacol. 132, 325–333.
Davis, S.F., Nation, J.R. and Mayleben, M.A. (1993) “The effects of chronic lead exposure on reactivity to frustrative nonreward in rats”,Toxicol. Lett. 66, 237–246.
Guo, T.L., Miller, M.A., Shapiro, I.M. and Shenker, B.J. (1998) “Mercuric chloride induces apoptosis in human T lymphocytes: evidence of mitochondrial dysfunction”,Toxicol. Appl. Pharmacol. 153, 250–257.
Hendershot, L., Wei, J., Gaut, J., Melnick, J., Aviel, S. and Argon, Y. (1996) “Inhibition of immunoglobulin folding and secretion by dominant negative BiP ATPase mutants”,Proc. Natl. Acad. Sci. USA 93, 5269–5274.
Holtzman, D., DeVries, C, Nguyen, H., Olson, J. and Bensch, K. (1984) “Maturation of resistance to lead encephalopathy: cellular and subcellular mechanisms”,Neurotoxicology 5, 97–124.
Hori, O., Matsumoto, M., Kuwabara, K., Maeda, Y, Ueda, H., Ohtsuki, T., Kinoshita, T., Ogawa, S., Stern, D.M. and Kamada, T. (1996) “Exposure of astrocytes to hypoxia/r-eoxygenation enhances expression of glucose-regulated protein 78 facilitating astrocyte release of the neuroprotective cytokine interleukin 6”,J. Neurochem. 66, 973–979.
Lazo, P.S., Dorfman, K., Noguchi, T., Mattei, M.G. and Bravo, R. (1992) “Structure and mapping of the fosB gene. FosB downregulates the activity of the fosB promoter”,Nucl. Acids Res. 20, 343–350.
Leborgne-Castel, N., Jelitto-Van Dooren, E.P., Crofts, A.J. and Denecke, J. (1999) “Overexpression of BiP in tobacco alleviates endoplasmic reticulum stress”,Plant Cell 11, 459–470.
Legare, M.E., Barhoumi, R., Burghardt, R.C. and Tiffany-Castiglioni, E. (1993) “Low-level lead exposure in cultured astroglia: identification of cellular targets with vital fluorescent probes”,Neurotoxicology 14, 267–272.
Legare, M.E., Barhoumi, R., Hebert, E., Bratton, G.R., Burghardt, R.C. and Tiffany-Castiglioni, E. (1998) “Analysis of Pb2+ entry into cultured astroglia”,Toxicol. Sci. 40, 90–100.
Lindahl, L.S., Bird, L., Legare, M.E., Mikeska, G., Bratton, G.R. and Tiffany-Castiglioni, E. (1999) “Differential ability of astroglia and neuronal cells to accumulate lead: dependence on cell type and on degree of differentiation”,Toxicol. Sci. 50, 236–243.
Lindquist, S. and Craig, E.A. (1988) “The heat-shock proteins”,Annu. Rev. Genet. 22, 631–677.
Little, E., Ramakrishnan, M., Roy, B., Gazit, G. and Lee, A.S. (1994) “The glucose-regulated proteins (GRP78 and GRP94): functions, gene regulation and applications”,Crit. Rev. Eukaryot. Gene. Expr. 4, 1–18.
Liu, H., Miller, E., van de Water, B. and Stevens, J.L. (1998) “Endoplasmic reticulum stress proteins block oxidant-induced Ca2+ increases and cell death”,J. Biol. Chem. 273, 12858–12862.
Miyake, H., Hara, I., Arakawa, S. and Kamidono, S. (2000) “Stress protein GRP78 prevents apoptosis induced by calcium ionophore, ionomycin, but not by glycosylation inhibitor, tunicamycin, in human prostate cancer cells”,J. Cell. Biochem. 77, 396–408.
Needleman, H.L., Riess, J.A., Tobin, M.J., Biesecker, G.E. and Greenhouse, J.B. (1996) “Bone lead levels and delinquent behavior”,J. Am. Med. Assoc. 275, 363–369.
Pishak, M.R. and Phillips, A.T. (1980) “Glucocorticoid stimulation of glutamine synthetase production in cultured rat glioma cells”,J. Neurochem. 34, 866–872.
Qian, Y., Tiffany-Castiglioni, E. and Harris, E.D. (1995) “Copper transport and kinetics in cultured C6 rat glioma cells”,Am. J. Physiol. 269, C892-C898.
Qian, Y., Tiffany-Castiglioni, E. and Harris, E.D. (1997) “A Menkes P-type ATPase involved in copper homeostasis in the central nervous system of the rat”,Mol. Brain Res. 48, 60–66.
Qian, Y, Tiffany-Castiglioni, E., Welsh, J. and Harris, E.D. (1998) “Copper efflux from murine microvascular cells requires expression of the Menkes disease Cu-ATPase”,J. Nutr. 128, 1276–1282.
Qian, Y., Mikeska, G., Harris, E.D., Bratton, G.R. and Tiffany-Castiglioni, E. (1999) “Effect of lead exposure and accumulation on copper homeostasis in cultured C6 rat glioma cells”,Toxicol. Appl. Pharmacol. 158, 41–49.
Qian, Y, Harris, E.D., Zheng, Y and Tiffany-Castiglioni, E. (2000) “Lead (Pb) targets GRP78, a molecular chaperone, in C6 rat glioma cells”,Toxicol. Appl. Pharmacol. 163, 260–266.
Ramesh, G.T., Manna, S.K., Aggarwal, B.B. and Jadhav, A.L. (1999) “Lead activates nuclear transcription factor-kappaB, activator protein-1 and ammo-terminal c-Jun kinase in pheochromocytoma cells”,Toxicol. Appl. Pharmacol. 155, 280–286.
Resendez, Jr., E., Wooden, Jr., S.K. and Lee, Jr., A.S. (1988) “Identification of highly conserved regulatory domains and protein-binding sites in the promoters of the rat and human genes encoding the stress-inducible 78-kilodalton glucose-regulated protein”,Mol. Cell. Biol. 8, 4579–4584.
Roy, B., Li, W.W and Lee, A.S. (1996) “Calcium-sensitive transcriptional activation of the proximal CCAAT regulatory element of the grp78/BiP promoter by the human nuclear factor CBF/NF-Y”,J. Biol. Chem. 271, 28995–29002.
Rushmore, T.H., Morton, M.R. and Pickett, C.B. (1991) “The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity”,J. Biol. Chem. 266, 11632–11639.
Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989) Molecular cloning: A laboratory manual, 2nd ed. (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY).
Sidrauski, C, Chapman, R. and Walter, P. (1998) “The unfolded protein response: An intracellular signalling pathway with many surprising features”,Trends Cell. Biol. 8, 245–249.
Tiffany-Castiglioni, E. (1993) “Cell culture models for lead toxicity in neuronal and glial cells”,Neurotoxicology 14, 513–536.
Tiffany-Castiglioni, E., Zmudzki, J., Wu, J.N. and Bratton, G.R. (1987) “Effects of lead treatment on intracellular iron and copper concentrations in cultured astroglia”,Metab. Brain Dis. 2, 61–79.
Tiffany-Castiglioni, E., Garcia, D.M., Wu, J.N., Zmudzki, J. and Bratton, G.R. (1988) “Effects of lead on viability and intracellular metal content of C6 rat glioma cells”,J. Toxicol. Environ. Health 23, 267–279.
Tiffany-Castiglioni, E., Legare, M.E., Schneider, L.A., Hanne-man, W.H., Zenger, E. and Hong, S.J. (1996) “Astroglia and lead neurotoxicity”, In: Aschner, M. and Kimelberg, K.H., eds, The Role of Glia in Neurotoxicity (CRC Press, Boca Raton, FL), pp 175–200.
Winder, C. (1984) The developmental neurotoxicity of lead (MTP Press, Lancaster).
Xiao, N. and DeFranco, D.B. (1997) “Overexpression of unliganded steroid receptors activates endogenous heat shock factor”,Mol. Endocrinol. 11, 1365–1374.
Xiao, G., Chung, T.F., Pyun, H.Y., Fine, R.E. and Johnson, R.J. (1999) “KDEL proteins are found on the surface of NG108-15 cells”,Brain Res. Mol. Brain Res. 72, 121–128.
Yu, Z.F., Luo, H., Fu, W. and Mattson, M.R (1999) “The endoplasmic reticulum stress-responsive protein GRP78 protects neurons against excitotoxicity and apoptosis: Suppression of oxidative stress and stabilization of calcium homeostasis”,Exp. Neurol. 155, 302–314.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Qian, Y., Falahatpisheh, M.H., Zheng, Y. et al. Induction of 78 kD glucose-regulated protein (GRP78) expression and redox-regulated transcription factor activity by lead and mercury in C6 rat glioma cells. neurotox res 3, 581–589 (2001). https://doi.org/10.1007/BF03033212
Received:
Revised:
Issue Date:
DOI: https://doi.org/10.1007/BF03033212