Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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The immune system has been identified as a sensitive target for the toxic effects produced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Furthermore, the B cell has been identified as a sensitive cellular target of TCDD by previous cell-type fractionation studies from this laboratory. The mechanism responsible for the immunotoxic effects produced by TCDD is unclear; however, many of the biological effects of TCDD are thought to be mediated by the aryl hydrocarbon receptor (AhR). Here, we describe two B cell lines that differ considerably in their expression of the AhR and in their sensitivity to TCDD. Our results demonstrated a marked expression of the AhR protein in the CH12.LX B cell line but not in the BCL-1 B cell line. Transcripts for the AhR were not detected by reverse transcriptase-polymerase chain reaction in the BCL-1 cells. The AhR nuclear translocator (ARNT) protein was highly expressed in both cell lines. In addition, the AhR and ARNT are functional in CH12.LX cells as demonstrated by TCDD-induced CYP1A1 induction. TCDD did not induce CYP1A1 in BCL-1 cells. Furthermore, TCDD treatment resulted in suppression of lipopolysaccharide (LPS)-induced IgM secretion in CH12.LX cells. Conversely, TCDD-induced inhibition of IgM secretion was not demonstrated in LPS-stimulated BCL-1 cells, implicating a role for the AhR in the inhibition of B cell effector function. LPS-induced differentiation of the CH12.LX cells also resulted in a marked induction of Ahr expression which was not induced in LPS-stimulated BCL-1 cells. These studies have implicated the AhR as a critical factor in TCDD-induced inhibition of IgM secretion and have demonstrated an induction of AhR gene and protein expression after B cell activation.
Mol Pharmacol 1998 Apr
PMID:Aryl hydrocarbon receptor-dependent suppression by 2,3,7, 8-tetrachlorodibenzo-p-dioxin of IgM secretion in activated B cells. 954 51

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most potent dioxin. There are exceptionally wide inter- and intraspecies differences in sensitivity to TCDD toxicity with Han/Wistar (H/W) (Kuopio) rats being the most resistant mammals tested. A peculiar feature of H/W rats is that despite their unresponsiveness to the acute lethality of TCDD, their sensitivity to other biological impacts of TCDD (e.g., CYP1A1 induction) is preserved. The biological effects of TCDD are mediated by the aryl hydrocarbon receptor (AhR). We recently found that the AhR of H/W rats (about 98 kDa) is smaller than the receptor in other rat strains (106 kDa). In the present study, molecular cloning and sequencing of the H/W rat AhR revealed that the reason for its smaller size is a deletion/insertion-type change at the 3' end of exon 10 in the receptor cDNA. This change emanates from a single point mutation at the first nucleotide of intron 10, resulting in altered mRNA splicing. At the protein level, the mutation leads to a total loss of either 43 or 38 amino acids (with altered sequence for the last seven amino acids in the latter case) toward the carboxyl-terminal end in the trans-activation domain of the AhR. H/W rats also harbor a point mutation in exon 10 that will cause a Val-to-Ala substitution in codon 497, but this occurs in a variable region of the AhR. These findings suggest that there is a relatively small region in the AhR trans-activation domain that may be capable of providing selectivity to its function.
Mol Pharmacol 1998 Jul
PMID:Point mutation in intron sequence causes altered carboxyl-terminal structure in the aryl hydrocarbon receptor of the most 2,3,7,8-tetrachlorodibenzo-p-dioxin-resistant rat strain. 965 93

A well-characterized primary rat hepatocyte culture system was used to examine induction patterns of cytochrome 450 gene expression by a series of 4-n-alkyl-methylenedioxybenzene (MDBs) derivatives. Hepatocytes were treated for 24, 48, or 72 hours with 0-500 microM of the MDB compounds, and total cellular RNA and protein from each treatment was evaluated by hybridization and immunochemical techniques. Exposure to MDB congeners possessing increasing 4-n-alkyl side-chain length (C0-C8) resulted in dose- and structure-dependent activation of CYP2B1, 2B2, 3A1, 1A1, and 1A2 gene expression. At equivalent 100 microM concentrations, the C6 and C8 MDB congeners were more effective than the prototypical inducer phenobarbital (PB) with respect to induction potency of CYP2B1, CYP2B2, and CYP3A1 gene expression. In contrast to PB, longer side-chain-substituted MDBs effectively induced CYP1A1 and CYP1A2 gene expression, in addition to the CYP2B and CYP3A genes. At equivalent molar concentrations, the catechol derivative of C6-MDB was ineffective in its ability to induce CYP gene expression, indicating the importance of the intact methylenedioxy bridge in the induction mechanism. Levels of MDB-inducible CYP2B1 and CYP2B2 mRNA were highly correlated with CYP2B1/2 apoprotein levels, ascertained by immunoblot analysis of cultured hepatocyte S9 fractions. Compared with results from previous in vivo analysis (12), the current data indicate that pharmacodynamic factors may influence MDB induction profiles and that differences in MDB effects on CYP gene expression result depending on distinct structure-activity relationships.
J Biochem Mol Toxicol 1998
PMID:Differential induction of cytochrome P450 gene expression by 4n-alkyl-methylenedioxybenzenes in primary rat hepatocyte cultures. 966 31

The effects of estradiol and progesterone on the expression of cytochrome P4501A1 were investigated in Hepa 1c1c7 cells. Both steroids, at 10 microM concentration, increased P4501A1-mediated 7-ethoxyresorufin O-deethyalase activity and amounts of its immunoreactive protein and CYP1A1 mRNA. Gel shift assay revealed that the steroids could induce both AhR transformation and binding of the ligand-AhR complex to its specific DNA recognition site. Transient transfection demonstrated that 5'flanking region of CYP1A1 could respond to the steroid action. The competitive binding assay showed that the steroids bound to AhR with moderate affinity. These results suggested that steroidal structure can be AhR ligands and induce CYP1A1 expression in AhR-dependent manner.
Biochem Mol Biol Int 1998 Jul
PMID:Effects of estradiol and progesterone on cytochrome P4501A1 expression in Hepa 1c1c7 cells. 971 1

Alterations in nutritional status affect hepatic cytochrome P450 levels. Since cytochromes P450 participate in the metabolism of arachidonic acid, we hypothesized that changes in liver P450 arachidonic acid metabolism occur during fasting and refeeding. Male Fisher 344 rats were either fed, fasted 48 hr (F48), fasted 48 hr and then refed 6 hr (F48/R6), or fasted 48 hr and then refed 24 hr (F48/R24). F48 rats had reduced body weight, increased plasma beta-hydroxybutyrate, and reduced plasma insulin compared with the other groups. Although there was no significant change in total liver P450 content, there was a significant 20%, 48%, and 24% reduction in total hepatic microsomal arachidonic acid metabolism in F48, F48/R6, and F48/R24 rats, respectively, compared with fed rats. Epoxygenase activity decreased by 28%, 51%, and 26% in F48, F48/R6, and F48/R24 rats, respectively. In contrast, omega-1 hydroxylase activity increased by 126% in F48 rats compared with fed rats. Immunoblotting revealed that levels of CYP2C11 protein were markedly reduced, whereas levels of CYP2E1 protein were markedly increased in the F48 and F48/R6 groups. In contrast, levels of CYP1A1, CYP1A2, CYP2B1, CYP2J3, CYP4A1, and CYP4A3 were unchanged with fasting/refeeding. Northern blots revealed that levels of CYP2C11 mRNAs were decreased, whereas CYP2E1 mRNAs were increased in F48 and F48/R6 rats. Recombinant CYP2C11 metabolized arachidonic acid primarily to epoxides with preference for the 14(S),15(R)-, 11(R), 12(S)-, and 8(S),9(R)- epoxyeicosatrienoic acid enantiomers. We conclude that (1) nutritional status affects hepatic microsomal arachidonic acid metabolism, (2) reduced epoxygenase activity in F48 and F48/R6 rats is accompanied by decreased levels of CYP2C11, (3) increased omega-1 hydroxylase activity is accompanied by augmented levels of CYP2E1, and (4) the effects of fasting on CYP2C11 and CYP2E1 expression occur at the pretranslational level.
Mol Pharmacol 1998 Sep
PMID:Nutritional status modulates rat liver cytochrome P450 arachidonic acid metabolism. 973 Sep 9

Our earlier investigation demonstrated that methoxychlor treatment induced hepatic cytochrome P450 2B1/2B2 and 3A proteins in rats and enhanced their respective enzymatic activities. To determine the mechanism of the methoxychlor-mediated induction and whether methoxychlor acts as a frank inducer and not merely by causing stabilization of the cytochrome P450 proteins or of their mRNAs, we assessed the effect of methoxychlor treatment on the transcriptional rate and mRNA levels of these hemeproteins by nuclear run-on and northern blot analyses. Methoxychlor treatment markedly elevated the transcriptional rates of cytochrome P4502B1/2B2 and 3A. Also, higher levels of mRNA of these CYPs were observed. By contrast, mRNA levels of CYP1A1/1A2 were not affected. Using a nuclear run-on method that apparently measures specifically the transcription initiation rate of CYP2B1 and 2B2, we observed that the transcriptional rate of 2B2 was increased substantially more than that of CYP2B1. This differential induction of CYP2B2 versus 2B1 may account for the discrepancy between the levels of induced CYP2B protein and the much lower-than-expected CYP2B-mediated enzymatic activity observed in our earlier study.
J Biochem Mol Toxicol 1998
PMID:Mechanism of induction of rat hepatic CYP2B and 3A by the pesticide methoxychlor. 973 80

Effects of chronic exposure to PCBs on the microsomal cytochrome P450 (CYP) enzymes in liver and testis of bulls (Bos taurus) were determined by comparing the constitutive and PCB-induced alkoxyresorufin O-dealkylase and testosterone hydroxylase activities. Specific inductions of the prevailing hepatic ethoxyresorufin O-deethylation and 6 beta-hydroxylation of testosterone are suggestive of the induction of CYP1A1 and CYP3A-like enzymes by PCBs. A high level of PCB-inducible androstenedione formation was also found. The hepatic CYP2B activities (i.e. pentoxyresorufin O-depentylase and testosterone 16 beta-hydroxylase) and CYP2C11-like testosterone 2 alpha-hydroxylase were increased only weakly. The testicular microsomal CYP activities were non-specifically reduced by the PCB exposure, except for the androstenedione formation and 16 beta-hydroxylation of testosterone. The inhibition of the activity of mitochondrial CYP11A, as the rate-limiting enzyme of steroidogenesis measured with resorufin 3 beta-hydroxy-22,23-bisnor-5-cholenyl ether as the fluorogenic substrate, exceeded 50% in testes of the PCB-contaminated bulls. The latter activity as well as the hepatic testosterone 6 beta-hydroxylation and hepatic and testicular androstenedione formation may significantly contribute to the decrease in testosterone levels after the PCB intake.
Comp Biochem Physiol A Mol Integr Physiol 1998 May
PMID:Effects of chronic exposure to PCBs on cytochrome P450 systems and steroidogenesis in liver and testis of bulls (Bos taurus). 977 99

Activation of the CYP1A1 gene has been described to be mediated by the cytosolic Ah receptor (AhR) and a possible cooperative role of the 4S benzo(a)pyrene-binding protein (4S protein). Carbaryl (CAR) has been shown to induce human CYP1A1 gene expression without binding to the human AhR. In this study, Sprague-Dawley rats received a single i.p. dose of 20, 80, 150 micromol/kg CAR or NAPn (naphthalene, the aromatic part of CAR) and were sacrificed after 24 h. CAR increased ethoxyresorufin-O-deethylase and methoxyresorufin-O-demethylase activities, the level of CYP1A1, 1A2 proteins, and CYP1A1 mRNA at the highest dose, whereas NAPn showed no effects. Moreover, CAR, naphthol (its major metabolite) and NAPn were not ligands in vitro of the TCDD binding site of AhR or the benzo(a)-pyrene binding site of 4S protein in rat, neither was CAR a ligand of these two binding sites in mice, dog, monkey or human. Molecular properties of CAR were evaluated and showed that this molecule is far from the structural characteristics of CYP 1A1 specific inducers although a planar conformation can be achieved with an energy < 5 kJ x mol(-1). The data demonstrated that CAR could also modulate the AhR-mediated responses, even though it did not meet the structural requirements to be ligand of AhR.
Int J Mol Med 1998 Nov
PMID:Effects of carbaryl and naphthalene on rat hepatic CYP1A1/2: potential binding to Ah receptor and 4S benzo(a)pyrene-binding protein. 985 62

Mice of the XVIInc/Z and DBA/2N strains, which are responsive and nonresponsive, respectively, to the aryl hydrocarbon (Ah) receptor, were treated with the hepatocarcinogen 5,9-dimethyldibenzo[c,g]carbazole and their livers were examined by nuclease P1-enhanced 32P-postlabeling for the levels of DNA adducts formed. Pretreatment at the doses usually reported in the literature with the cytochrome P4501A (CYP1A) inducers 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), beta-naphthoflavone (BNF), and isosafrole modulated DNA adduction. In XVIInc/Z mice, DNA adduction was totally inhibited by TCDD (a CYP1A1/1A2 inducer), BNF (a CYP1A1/1A2 inducer), and isosafrole (a CYP1A2 inducer). In DBA/2N mice, in which DNA adduction was also inhibited by TCDD, about 25% of the DNA adduct levels persisted after pretreatment with BNF (not a CYP1A1/1A2 inducer in this strain) or isosafrole (a CYP1A2 inducer in this strain). The increase (in all cases less than twofold) in the levels of the phase-II drug-metabolizing enzymes glutathione S-transferase and uridine diphospho-glucuronyltransferase after treatment with inducers cannot explain the total disappearance of DNA adducts. Assays of 5-bromo-2'-deoxyuridine incorporation did not show any induction of DNA synthesis which could explain the decrease in adducts. These results suggest that in vivo 1) increases in CYP1A enzymes by inducers are not correlated with enhanced levels of certain DNA adducts; and 2) phase-II drug metabolizing enzymes are not the main cellular protection pathway for detoxification. An additional mechanism, perhaps also induced by the Ah receptor but highly dependent on the dose of inducer, could be involved in parallel to multidrug resistance (mdr); further experiments are needed to identify this process used by the cell to enhance its protection against toxic or genotoxic effects.
Environ Mol Mutagen 1998
PMID:Inhibition of 5,9-dimethyldibenzo[c,g]carbazole-DNA adduct formation in mouse liver by pretreatment with cytochrome P4501A inducers in vivo. 988 5

In this investigation, we examined the effects of insulin on gene induction responsiveness in primary rat hepatocytes. Cells were cultured for 72 hours either in the absence or presence of 1 microM insulin and then exposed to increasing concentrations of phenobarbital (PB; 0.01-3.5 mM). Culturing in the absence of insulin produced 1.5-2-fold increases in the induction magnitude of CYP2B1 and CYP2B2 mRNA expression resulting from PB exposures, without altering the bell-shaped dose-response curve characteristic of this agent. However, for the CYP3A1 gene, insulin removal led to a pronounced shift in both the PB-induction magnitude and dose-response relationships of the induction response, with higher levels of CYP3A1 expression resulting from exposures to lower concentrations of inducer. Insulin removal also reduced the time required to attain maximal induction of CYP2B1/2 and CYP3A1 gene expression. The insulin effects were not specific for PB induction, as insulin deprivation similarly enhanced both dexamethasone- and beta-naphthoflavone-inducible CYP3A1 and CYP1A1 expression profiles, respectively. In contrast, the level of albumin mRNA expression was reduced considerably in cells deprived of insulin. We conclude that insulin is an important regulator of inducible and liver-specific gene expression in primary rat hepatocytes.
J Biochem Mol Toxicol 1999
PMID:Insulin-mediated modulation of cytochrome P450 gene induction profiles in primary rat hepatocyte cultures. 989 Apr 42


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