Overexpression of Metallothionein-II Sensitizes Rodent Cells to Apoptosis Induced by DNA Cross-linking Agent through Inhibition of NF-κB Activation

DNA cross-linking agents such as mitomycin C (MMC) and cisplatin are used as chemotherapeutic agents in cancer treatment. However, the molecular mechanism underlying their antitumor activity is not entirely clear. Critical steps in cytotoxicity toward cross-linking agents can involve DNA repair efficiency, inhibition of replication, cell-cycle checkpoints, regulation, and induction of apoptosis. The complexity of the mechanisms of the mammalian cell defense against cross-linking agents is reflected by the existence of many complementation groups identified in rodent cells that are specifically sensitive to MMC. We recently showed that increased induction of apoptosis contributes to the MMC sensitivity of the group represented by the V-H4 hamster mutant cell line. In this study, through the analyses of a substractive library, we discovered that sensitive V-H4 cells display a 40-fold increase of steady-state expression of metallothionein II (MT-II) mRNA compared with resistant parental V79 cells. Down-regulation of MT-II by antisense oligonucleotides partially restores MMC resistance in V-H4 cells, indicating that MT-II overexpression is directly involved in MMC hypersensitivity of these cells. MTs have been reported to regulate the activation of NF-κB, one of the key proteins that modulates the apoptotic response. Here we found that NF-κB activation by MMC is impaired in V-H4 cells and is partially restored following down-regulation of MT-II by antisense oligonucleotides. All these data suggest that the overexpression of MT-II in V-H4 cells impairs NF-κB activation by MMC, resulting in decreased cell survival and enhanced induction of apoptosis. DNA cross-linking agents such as mitomycin C (MMC) and cisplatin are used as chemotherapeutic agents in cancer treatment. However, the molecular mechanism underlying their antitumor activity is not entirely clear. Critical steps in cytotoxicity toward cross-linking agents can involve DNA repair efficiency, inhibition of replication, cell-cycle checkpoints, regulation, and induction of apoptosis. The complexity of the mechanisms of the mammalian cell defense against cross-linking agents is reflected by the existence of many complementation groups identified in rodent cells that are specifically sensitive to MMC. We recently showed that increased induction of apoptosis contributes to the MMC sensitivity of the group represented by the V-H4 hamster mutant cell line. In this study, through the analyses of a substractive library, we discovered that sensitive V-H4 cells display a 40-fold increase of steady-state expression of metallothionein II (MT-II) mRNA compared with resistant parental V79 cells. Down-regulation of MT-II by antisense oligonucleotides partially restores MMC resistance in V-H4 cells, indicating that MT-II overexpression is directly involved in MMC hypersensitivity of these cells. MTs have been reported to regulate the activation of NF-κB, one of the key proteins that modulates the apoptotic response. Here we found that NF-κB activation by MMC is impaired in V-H4 cells and is partially restored following down-regulation of MT-II by antisense oligonucleotides. All these data suggest that the overexpression of MT-II in V-H4 cells impairs NF-κB activation by MMC, resulting in decreased cell survival and enhanced induction of apoptosis. DNA cross-linking agents constitute a set of pharmacological molecules used as single agents or in combination in the treatment of a wide variety of malignant tumors (1Kohn K.W. Cancer Res. 1996; 56: 5533-5546PubMed Google Scholar). Among them, mitomycin C (MMC) 1The abbreviations used are:MMCmitomycin CMT-IImetallothionein-IITNFtumor necrosis factorCPTcamptothecinNFnuclear factorEMSAelectrophoretic mobility shift assay is more used particularly for the treatment of adenocarcinomas, non-small cell lung cancer, some head and neck cancers, and in chronic myelogenous leukemia (2Sartorelli A.C. Hodnick W.F. Belcourt M.F. Tomasz M. Haffty B. Fischer J.J. Rockwell S. Oncol. Res. 1994; 6: 501-508PubMed Google Scholar). MMC is activated in vivo to an alkylating agent by a reductive activation cascade and forms monoadducts and interstrand or intrastrand cross-links on DNA, preferentially at the N2 position of guanine (3Tomasz M. Palom Y. Pharmacol. Ther. 1997; 76: 73-87Crossref PubMed Scopus (240) Google Scholar). The molecular mechanism underlying the MMC antitumor activity is not fully understood. To elucidate the mechanisms of the mammalian cell defense against cross-linking agents, mutants specifically sensitive to MMC have been isolated in rodent cells (4Zdzienicka M.Z. Simons J.W. Mutat. Res. 1987; 178: 235-244Crossref PubMed Scopus (96) Google Scholar). The genetic and biochemical complexity of these processes is reflected by the existence of at least eight complementation groups identified among rodent cell mutants defective in the response to MMC treatment (5Telleman P. Overkamp W.J. van Wessel N. Studzian K. Wetselaar L. Natarajan A.T. Zdzienicka M.Z. Cancer Res. 1995; 55: 3412-3416PubMed Google Scholar, 6Jones N.J. Mutagenesis. 1994; 9: 477-482Crossref PubMed Scopus (25) Google Scholar). mitomycin C metallothionein-II tumor necrosis factor camptothecin nuclear factor electrophoretic mobility shift assay V-H4 cell line, a representative of one of these complementation group, was isolated from V79 Chinese hamster cells (7Zdzienicka M.Z. Arwert F. Neuteboom I. Rooimans M. Simons J.W. Somatic Cell Mol. Genet. 1990; 16: 575-581Crossref PubMed Scopus (31) Google Scholar). V-H4 mutant cells exhibit increased sensitivity toward cross-linking agents such as MMC (∼30-fold more sensitive than the wild-type V79 cells) and cisplatin (∼10-fold more sensitive), but they are not hypersensitive to UV light, H2O2, or x-rays (7Zdzienicka M.Z. Arwert F. Neuteboom I. Rooimans M. Simons J.W. Somatic Cell Mol. Genet. 1990; 16: 575-581Crossref PubMed Scopus (31) Google Scholar). The V-H4 cell response to this panel of genotoxic agents suggests that the defective protein or pathway in these cells is specific to cross-linking agents. However, the molecular defect responsible for the sensitivity of this mutant cell line to MMC remains to be determined. Critical steps in cytotoxicity toward cross-linking agents can involve DNA repair efficiency (8Larminat F. Bohr V.A. Nucleic Acids Res. 1994; 22: 3005-3010Crossref PubMed Scopus (60) Google Scholar), inhibition of replication (9Plooy A.C. van Dijk M. Berends F. Lohman P.H. Cancer Res. 1985; 45: 4178-4184PubMed Google Scholar), cell-cycle checkpoints regulation, and induction of programmed cell death (10Demarcq C. Bunch R.T. Creswell D. Eastman A. Cell Growth Differ. 1994; 5: 983-993PubMed Google Scholar). We showed previously that neither a defect in nucleotide excision repair of DNA interstrand cross-links (11Larminat F. Cambois G. Zdzienicka M.Z. Defais M. FEBS Lett. 1998; 437: 97-100Crossref PubMed Scopus (11) Google Scholar) nor a defective G2 phase checkpoint contributes to the differential sensitivity of V-H4 mutant toward MMC (12Papouli E. Lafon C. Valette A. Zdzienicka M.Z. Defais M. Larminat F. Biochem. Pharmacol. 2000; 59: 1101-1107Crossref PubMed Scopus (10) Google Scholar). In contrast, our findings demonstrated that sensitive V-H4 cells undergo greater levels of apoptosis than resistant parental cells following both equimolar and equitoxic MMC treatment (12Papouli E. Lafon C. Valette A. Zdzienicka M.Z. Defais M. Larminat F. Biochem. Pharmacol. 2000; 59: 1101-1107Crossref PubMed Scopus (10) Google Scholar). This differential apoptotic response is specific for the cross-linking agent MMC and is p53-independent, because p53 sequence is mutated in V-H4 cells (12Papouli E. Lafon C. Valette A. Zdzienicka M.Z. Defais M. Larminat F. Biochem. Pharmacol. 2000; 59: 1101-1107Crossref PubMed Scopus (10) Google Scholar) as well as in V79 parental cells (13Chaung W., Mi, L.J. Boorstein R.J. Nucleic Acids Res. 1997; 25: 992-994Crossref PubMed Scopus (115) Google Scholar). Our previous results suggested then that control of the apoptotic process is altered in V-H4 mutant cells. Defective gene(s) in these cells could function in the regulation of an apoptotic pathway triggered by MMC-induced damages and independent of p53-mediated transcription. Product(s) of this gene(s) could interfere at different levels of this process, such as detection of the MMC adduct on the DNA or of an intermediate of lesion repair, presence of reactive oxygen species produced during MMC detoxification, or even being directly implicated in the transduction cascade of the apoptotic signal initiated by MMC. To further characterize the molecular defect in V-H4 cell line, we sought to identify gene(s) involved in MMC hypersensitivity of these cells. In this report, we compared mRNA expression pattern of V-H4 mutant cell line with that of V79 parental cell line. Using a suppression substractive hybridization methodology (14Diatchenko L. Lau Y.F. Campbell A.P. Chenchik A. Moqadam F. Huang B. Lukyanov S. Lukyanov K. Gurskaya N. Sverdlov E.D. Siebert P.D. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 6025-6030Crossref PubMed Scopus (2748) Google Scholar), we found a steady-state overexpression of metallothionein-II (MT-II) gene in V-H4 mutant cell line. MT-II is a small cystein-rich, heavy metal-binding protein that participates in detoxification pathways and regulation of cell homeostasis (15Palmiter R.D. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8428-8430Crossref PubMed Scopus (612) Google Scholar). The increased level of MT-II protein in the MMC-hypersensitive V-H4 cells was an unexpected finding, because MT-II overexpression is generally associated with drug resistance (16Kaina B. Lohrer H. Karin M. Herrlich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 2710-2714Crossref PubMed Scopus (97) Google Scholar, 17Kelley S.L. Basu A. Teicher B.A. Hacker M.P. Hamer D.H. Lazo J.S. Science. 1988; 241: 1813-1815Crossref PubMed Scopus (610) Google Scholar). In this study, we have investigated the role of MT-II overexpression in MMC hypersensitivity of V-H4 cell line and report for the first time that MT-II overexpression can sensitize hamster cells to apoptosis induced by DNA cross-linking agent through NF-κB inhibition. The MMC-sensitive mutant V-H4 was previously isolated from Chinese hamster V79 cells and was kindly provided by Professor M. Z. Zdzienicka (7Zdzienicka M.Z. Arwert F. Neuteboom I. Rooimans M. Simons J.W. Somatic Cell Mol. Genet. 1990; 16: 575-581Crossref PubMed Scopus (31) Google Scholar). V79 and V-H4 cell lines were routinely grown in monolayer in Ham's F-10 medium (Invitrogen) supplemented with 15% newborn calf serum (Invitrogen), penicillin (100 units/ml), and streptomycin (0.1 μg/ml). All incubations were at 37 °C in humidified 5% CO2 atmosphere. Cells were tested by PCR for mycoplasma prior to suppression substractive hybridization methodology experiments. For all experiments in this study, cells were in exponential growth phase. Cells were treated with MMC (Sigma) or TNFα (Chemicon) for 1 h, cadmium chloride (Sigma) for 2 h, or camptothecin (CPT) (Sigma) for 3 h at the indicated concentrations at 37 °C. After drug exposure, cells were washed with and then with mRNA was the substractive hybridization methodology was the were by PCR and the at was of were in and to with at 37 °C in of medium were then a set a and were grown at 37 °C. were by a first with for and then with for DNA was on by for at °C. After at °C with were with for h at °C. were by DNA and The was with and and the as in the were washed in at °C for and in at °C for were a and the After DNA were identified the at the were isolated the (Invitrogen) to the from the were to in and then were with a MT-II to the mRNA sequence of Chinese hamster MT-II mRNA for h at °C. were a and the MT-II mRNA was by mRNA to cadmium or MMC was in a growth Cells were at h prior treatment to cells were then treated with different concentrations of 2 h or MMC for 1 After drug cells were with and medium was Cells were to for and were then and in and were in at Cell survival was to the of of V79 and V-H4 cells were routinely and oligonucleotides were an antisense sequence and a sequence to the from the of MT-II mRNA sequence and The sequence to a to the of the a resistant to B. G. B. 1995; 6: PubMed Scopus Google Scholar). cells were in of and to 1 of 1 cells were in Ham's F-10 medium supplemented with 15% newborn calf serum and at of the antisense in MT-II protein was by decreased sensitivity to cadmium a growth assay h with The of MT-II down-regulation on MMC sensitivity was h with antisense were from or cells as previously Nucleic Acids Res. PubMed Scopus Google Scholar) with a Cells were in of supplemented by to on for for and for 1 at were in of C supplemented by and on for for proteins were the of by for 2 at were with the NF-κB sequence and 1 of (Sigma) in 1 for at °C. DNA were by both of the and of was used as a control to specifically for mutated was used to the of of NF-κB to the DNA not were a were in at for 1 h, and to To a differential gene expression that be involved in MMC hypersensitivity of V-H4 mutant cells, we compared the steady-state mRNA expression parental V79 and V-H4 mutant cell Using suppression substractive we isolated a a to the sequence of hamster metallothionein-II of MT-II mRNA in V-H4 cells was by hybridization of to the to MT-II as on the representative in increase of steady-state MT-II gene expression was found in mutant V-H4 cells compared with the parental cell line V79 results were with independent MT-II is induced by and agents such as growth and S.L. L. Nucleic Res. Mol. 1998; 59: PubMed Scopus Google Scholar). We investigated MT-II mRNA levels can be increased by MMC treatment. V79 and V-H4 cells were treated with MMC for 1 h, and MT-II expression was at drug treatment. Our results showed that MT-II mRNA is induced as as treatment in V79 cell line induction steady-state in V-H4 cells to a 1 h induction that MT-II mRNA remains in response to in V-H4 cells level In both cell MT-II mRNA levels at 3 h MMC treatment to levels and decreased by h We then MT-II mRNA overexpression in V-H4 cells was associated with increased MT-II protein been that in mRNA for well with the induction of protein R.D. PubMed Google Scholar). levels could not be in our cell lines by or as previously Mol. Pharmacol. Google Scholar), because against with hamster MT-II However, a been levels and resistance to S. Pharmacol. PubMed Scopus Google Scholar). can heavy such as or cadmium and cells against their cytotoxicity (15Palmiter R.D. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 8428-8430Crossref PubMed Scopus (612) Google Scholar). been reported that the overexpression of the MT-II gene in Chinese hamster cell lines to a resistance to of these cells Mol. Pharmacol. Google Scholar, A. Lohrer H. Mutat. Res. PubMed Scopus Google Scholar). we cell survival to to MT-II mRNA overexpression in V-H4 cells is associated with increased MT-II of V79 and V-H4 cells demonstrated that V-H4 cells are more resistant to than parental cells, the to cell survival to of V79 and V-H4 being and results that V-H4 cells levels of MT-II proteins than the parental V79 cells. To MT-II overexpression was directly involved in MMC hypersensitivity of V-H4 mutant cells, we used antisense oligonucleotides to MT-II protein Cells were with a antisense the of MT-II mRNA or with the antisense oligonucleotides were previously used to MT-II protein expression in V79 cells M. PubMed Scopus Google Scholar). MT-II expression by antisense oligonucleotides was by assay following treatment 3 After with the antisense MT-II V-H4 as sensitive as V79 cells to because was decreased from to on V-H4 survival results that antisense MT-II oligonucleotides decreased the MT-II protein in V-H4 cells. We then investigated the of MT-II down-regulation on MMC resistance of the mutant cells. After with V-H4 cells were treated for 1 h with of MMC as V-H4 cell survival showed with or treatment with the oligonucleotides 3 In contrast, a of V-H4 cells with antisense oligonucleotides increased cell resistance to MMC, because increased from to results that MT-II overexpression at least in to MMC hypersensitivity of V-H4 mutant cells. The role of MT-II in detoxification processes by molecules is well and the increase of levels of metallothionein been associated with the of resistance to the of some alkylating agents (16Kaina B. Lohrer H. Karin M. Herrlich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 2710-2714Crossref PubMed Scopus (97) Google Scholar). However, role of MT-II in the regulation of processes following drug treatment been reported Mol. Pharmacol. Google Scholar, A. Lohrer H. Mutat. Res. PubMed Scopus Google Scholar). data have suggested a role of MT-II in the regulation of NF-κB, a factor A. S. N. Y. N. FEBS Lett. PubMed Scopus Google Scholar). The of are activated by a wide of DNA and D. 1996; 87: PubMed Scopus Google Scholar). In cells, NF-κB is in an in the by with of the such as and S. Cancer Res. 1995; PubMed Google Scholar). is on the protein for and by the S. 1995; PubMed Scopus Google Scholar). NF-κB is then to the and the of a variety of cell as well as proteins Science. 1998; PubMed Scopus Google Scholar). MT-II the activation of NF-κB and of an of implicated in cell survival and control of apoptosis Res. 437: PubMed Scopus Google Scholar). To further characterize the molecular pathway MT-II in to MMC treatment and overexpression can cell survival in V-H4 cells, we have NF-κB activation in these cells. The activation of NF-κB was MMC treatment by through to a specific DNA of was by with an of levels of steady-state activation of NF-κB were in both resistant and sensitive cell lines 1 and In wild-type V79 cells, was a activation of NF-κB following MMC following the of a drug and was increase of activity compared with cells) contrast, a NF-κB activation was in V-H4 mutant cells increase of We MT-II protein overexpression was involved in impaired NF-κB activation MMC treatment in V-H4 mutant cells. NF-κB activation was following MMC treatment in mutant cells with MT-II oligonucleotides Our results showed that MT-II protein down-regulation in a increase of NF-κB activity following MMC treatment in V-H4 cells 1 and findings that MT-II overexpression directly or NF-κB activation following MMC treatment in V-H4 cells. To the of this we NF-κB activation following TNFα is to this In V79 cells, a increase of NF-κB activity was 1 h TNFα and this activation to h In V-H4 cells, we a NF-κB activation following TNFα treatment NF-κB activation was h following TNFα treatment in mutant cells To elucidate the impaired activation of NF-κB by MMC and TNFα was because of a to NF-κB in sensitive cells, we NF-κB activity following camptothecin treatment and of in DNA of the with a replication or a and to death Mol. Pharmacol. 1996; Google Scholar). Our results showed a and increase of NF-κB activity in both cell lines by treatment that NF-κB activation is not defective in V-H4 mutant cells. our findings suggest that MT-II protein overexpression NF-κB activation by MMC treatment in V-H4 cells. we an altered NF-κB activation in mutant cells following both MMC and TNFα that MMC this factor through a or partially pathway with this sensitivity to cross-linking agents such as MMC is a of the V-H4 hamster cell line (7Zdzienicka M.Z. Arwert F. Neuteboom I. Rooimans M. Simons J.W. Somatic Cell Mol. Genet. 1990; 16: 575-581Crossref PubMed Scopus (31) Google Scholar). We previously reported that an increased induction of apoptosis contributes to sensitivity toward DNA cross-linking agents (12Papouli E. Lafon C. Valette A. Zdzienicka M.Z. Defais M. Larminat F. Biochem. Pharmacol. 2000; 59: 1101-1107Crossref PubMed Scopus (10) Google Scholar). To identify gene(s) involved in the MMC hypersensitivity of these cells, we compared mRNA expression of parental V79 and sensitive V-H4 cells. In this report, we showed that V-H4 cells display a 40-fold increase of steady-state expression of MT-II mRNA compared with resistant parental V79 cells. This MT-II mRNA is with a MT-II protein overexpression as by the increased resistance of V-H4 cells toward we not MT-II is in these MMC-hypersensitive cells. The mechanism of MT-II regulation at the level of M. R.D. E. K. PubMed Scopus Google Scholar). In to a of of MT-II such as and S.L. L. Nucleic Res. Mol. 1998; 59: PubMed Scopus Google Scholar), a factor that the MT-II and MT-II gene been recently identified E. Mol. PubMed Scopus (25) Google Scholar). is that the level of protein or of MT-II is involved in the overexpression of MT-II that we found in V-H4 cells. of the of this protein as a of reactive molecules and against J.S. 1998; PubMed Scopus Google Scholar, D. D. C. Mol. 2000; Google Scholar). our that MT-II is in MMC-hypersensitive V-H4 cells was not with previous data an increased resistance to resulting from MT-II overexpression (16Kaina B. Lohrer H. Karin M. Herrlich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 2710-2714Crossref PubMed Scopus (97) Google S.L. Basu A. Teicher B.A. Hacker M.P. Hamer D.H. Lazo J.S. Science. 1988; 241: 1813-1815Crossref PubMed Scopus (610) Google Scholar). Our that MT-II not in this hamster mutant cell line as a for alkylating agents but is associated with the regulation of with this and our overexpression of the gene the control of a is reflected in decreased resistance to cross-linking agent in wild-type Chinese hamster cells Mol. Pharmacol. Google Scholar). the alkylating hamster cells the gene an increased sensitivity to and MMC A. Lohrer H. Mutat. Res. PubMed Scopus Google Scholar). these for a of overexpression on drug resistance in Chinese hamster cell However, in these was this sensitivity to cross-linking agents was a of or apoptotic cell death and proteins regulate drug To MT-II overexpression was directly involved in MMC hypersensitivity of V-H4 cell line, we used antisense oligonucleotides to MT-II protein The efficiency of these oligonucleotides was by a well assay that cell sensitivity to heavy such as cadmium and was previously used in hamster cell lines Mol. Pharmacol. Google Scholar, A. Lohrer H. Mutat. Res. PubMed Scopus Google Scholar) and particularly in V79 cells M. PubMed Scopus Google Scholar). V-H4 cells a increase in sensitivity following MT-II protein the antisense inhibition of MT-II is a of the resistance to MMC was in the mutant cell line. we that MT-II overexpression contributes to MMC hypersensitivity in V-H4 cells. We MT-II overexpression could be directly involved in MMC sensitivity by with cell been reported that regulate NF-κB activation A. S. N. Y. N. FEBS Lett. PubMed Scopus Google Scholar). NF-κB is a factor associated with particularly through the induction of an of Res. 437: PubMed Scopus Google Scholar). The of activation of this factor could the increased induction of apoptosis in V-H4 cells MMC treatment H. A. K. 2000; PubMed Scopus Google Scholar, Y. M. P. S. Cancer Res. Scopus (97) Google Scholar, A. B. Oncol. PubMed Scopus Google Scholar). Our results showed that DNA activity of NF-κB following MMC treatment is in V-H4 cells as compared with the activation in V79 parental cells. the NF-κB pathway induced by MMC treatment be to be in V-H4 cells. the NF-κB activation following treatment in both sensitive and resistant cells, that activation of NF-κB, this factor to the and to regulate is not defective in V-H4 cells. DNA by MMC and NF-κB through different pathways because of in the of DNA or because NF-κB activation not involve nuclear but a We compared the of TNFα to the NF-κB activation in mutant and parental cells. Our results showed that NF-κB activation TNFα treatment is in V-H4 mutant cells. We that MMC and TNFα NF-κB through a of that some through activation of cell death such as E. A. PubMed Scopus Google Scholar, Cell Res. 2000; PubMed Scopus Google Scholar, S. E. M. K. 93: PubMed Google Scholar). The activation of these death to cell survival by NF-κB activation through G. A. F. F. M. Biochem. Pharmacol. 1998; 56: PubMed Scopus Google Scholar) or apoptosis following and activation E. A. PubMed Scopus Google Scholar). NF-κB activation cell enhanced NF-κB activity cells from In with this of activation of NF-κB by MMC in V-H4 cells results in decreased cell survival and enhanced induction of apoptosis triggered by damages (12Papouli E. Lafon C. Valette A. Zdzienicka M.Z. Defais M. Larminat F. Biochem. Pharmacol. 2000; 59: 1101-1107Crossref PubMed Scopus (10) Google Scholar). can MT-II regulate NF-κB of NF-κB in response to or to and a of NF-κB to the S. 1995; PubMed Scopus Google Scholar). of proteins in the of cells at the time of the NF-κB could for MT-II function as a of is well that 2000; PubMed Google Scholar). increase of MT-II levels in V-H4 cells to by C. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: PubMed Scopus Google Scholar), and this been to in decreased NF-κB activation M.P. 2000; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, U. PubMed Google Scholar) and induction of apoptosis Biochem. Res. 2000; PubMed Scopus Google Scholar, P.D. Biochem. Res. 2000; PubMed Scopus Google Scholar). However, is to be to a to NF-κB in response to and we activation of NF-κB following treatment in both V79 and V-H4 cells. the a regulation of NF-κB through a with been with the of NF-κB and be to DNA of NF-κB treatment Cancer Res. 1998; Google M. 2000; PubMed Scopus Google Scholar). However, this suggest a of NF-κB by our results that MT-II overexpression to specific inhibition of NF-κB by MMC in V-H4 cells. one could that MT-II is involved in the regulation of a specific pathway that We showed that NF-κB activation in response to is different from that triggered by such as MMC and Our findings the existence of different transduction pathways in rodent cells on the of the NF-κB could be to pathways through the of different on the and the of in different cell of be then activated by a specific In our findings for a of MT-II overexpression to MMC hypersensitivity of hamster V-H4 cells through the inhibition of NF-κB activation and enhanced apoptotic of the role of MT-II and NF-κB be to the mechanism of cell death induced by DNA cross-linking agents. the mechanisms of MT-II function in inhibition of NF-κB to the of agents of tumor cells to or DNA apoptosis. We M. P. for in experiments.