Effect of dexamethasone and phenobarbital on run-on transcription rate and CYP3A mRNA concentration in rat liver: Changes during development

Telhada, Margarida; Pereira, Teresa; Lechner, Maria Celeste

doi:10.1016/0003-9861(92)90471-8

Cited by 43 publications

(27 citation statements)

References 40 publications

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“…Also shown aligned with CYP3A23 is the homologous region of the CYP3A2 gene. Within this region of the two promoters, the genes are 91% identical, but the promoters differ in the magnitude of their response to glucocorticoids (23). It is likely that mismatches between CYP3A23 and 3A2, indicated ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…CYP3A2 is the form expressed in uninduced animals; however, its expression displays a gender and developmental stage-dependent pattern (8, 20 -22). Despite its high homology (89%) to 3A23, CYP3A2 is only weakly induced by glucocorticoids (23,24). CYP3A23 is expressed at low levels in untreated animals but is highly inducible by glucocorticoids, such as dexamethasone, and by PCN, a synthetic steroid with no hormonal activity (16,25).…”

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confidence: 99%

“…CYP3A23 is expressed at low levels in untreated animals but is highly inducible by glucocorticoids, such as dexamethasone, and by PCN, a synthetic steroid with no hormonal activity (16,25). Results from nuclear run-on experiments have established that the response is transcriptional (23,26).…”

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Nuclear Receptor Involvement in the Regulation of Rat Cytochrome P450 3A23 Expression

Huss¹,

Kasper

1998

Journal of Biological Chemistry

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Many genes of the cytochrome P450 3A (CYP3A) subfamily, including several human and rat isoforms, are inducible by glucocorticoids. In the rat CYP3A23 gene, a 110-base pair segment of the proximal 5-flanking region mediates dexamethasone activation. Three binding sites (DexRE-1, DexRE-2, and Site A), identified by DNase I footprinting analysis, were characterized for their relative contribution to both basal activity and dexamethasone inducibility. Site-directed mutagenesis of DexRE-1 (؊144 to ؊169) and DexRE-2 (؊118 to ؊136) demonstrated that each contained a core imperfect AGGTCA direct repeat, which comprised a consensus nuclear receptor binding site, and was essential for dexamethasone responsiveness but was not required for basal activity. Competition gel shift and supershift analyses revealed that both sites can bind the orphan nuclear receptor chicken ovalbumin upstream promoter-transcription factor.Site A (؊85 to ؊110) was shown to be important for both basal activity and dexamethasone responsiveness. Point mutants displayed a reduced (2-3-fold) induction response, compared with 15-fold for wild-type, which was accompanied by a 40 -60% drop in basal activity. Site A was shown to bind the liver-enriched nuclear receptor hepatocyte nuclear factor 4. Our studies demonstrate that the mechanism mediating glucocorticoidinducible transcriptional activity of CYP3A23 involves multiple binding sites for members of the nuclear receptor superfamily.The cytochrome P450s (CYPs) 1 make up a superfamily of heme-containing monooxygenases that are found most abundantly in liver endoplasmic reticulum and catalyze the oxidation or reduction of both endogenous and foreign substrates, often as an essential step in their elimination (1). CYP expression is modulated by endogenous and exogenous compounds, which may reflect a homeostatic mechanism whereby normal "endogenous ligand" concentrations are maintained. In early studies, steroids were shown to regulate expression of a cytochrome P450 family that was distinct from the 3-methylcholanthrene or phenobarbital inducible P450s (2-5). Pregnenolone 16␣-carbonitrile (PCN) was characterized as the prototypical inducer of the CYP3A family; however, many glucocorticoids have been shown to be more potent than PCN, whereas certain non-glucocorticoids, such as phenobarbital and macrolide antibiotics, also induce the same protein (6 -8). CYP3A proteins have been identified in several species, including rat, rabbit, mouse, and human (9 -14). The major glucocorticoid-responsive CYP3A gene in rat is 3A23, whereas 3A4 and 3A5 are inducible forms in human (15, 16). CYP3A enzymes metabolize over 60% of therapeutically relevant compounds and are involved in 6␤-hydroxylation of the endogenous steroids testosterone, progesterone, and cortisol (17-19).Regulation of rat 3A genes CYP3A23 and 3A2 has been the most extensively characterized. CYP3A2 is the form expressed in uninduced animals; however, its expression displays a gender and developmental stage-dependent pattern (8, 20 -22). Despite its hi...

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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Nuclear Receptor Involvement in the Regulation of Rat Cytochrome P450 3A23 Expression

Huss¹,

Kasper

1998

Journal of Biological Chemistry

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“…All CYP3A genes have been reported to undergo induction by many drugs and environmental or dietary substances, and this phenomenon can lead to clinically relevant adverse drug interactions (Thummel and Wilkinson, 1998). In addition, since CYP3A genes appear to be regulated mostly at the transcription level (Telhada et al, 1992;Schuetz et al, 1993;Li et al, 1995b), mRNA quantification should give a fair estimate of the variability of their expression. Interestingly, some, albeit not all, workers in the field have reported the expression and even the induction of CYP3A genes in peripheral blood leukocytes (Janardan et al, 1996;Hukkanen et al, 1997;Sempoux et al, 1999;Starkel et al, 1999;Krovat et al, 2000;Nakamoto et al, 2000;Asghar et al, 2002;Finnstrom et al, 2002;Nowakowski-Gashaw et al, 2002).…”

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confidence: 99%

Interindividual Variability and Tissue-Specificity in the Expression of Cytochrome P450 3A mRNA

Koch¹,

Weil²,

Wolbold³

et al. 2002

Drug Metab Dispos

280

188

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ABSTRACT:The elucidation of the individual contributions of the four CYP3A genes to the overall CYP3A activity has been hampered by similarities in their sequence and function. We investigated the expression of CYP3A mRNA species in the liver and in various other tissues using gene-specific TaqMan probes. CYP3A4 transcripts were the most abundant CYP3A mRNA in each of the 63 white European livers tested and accounted on average for 95% of the combined CYP3A mRNA pool. CYP3A5 and CYP3A7 each contributed on average 2%, whereas CYP3A43 contributed 0.3% transcripts to this pool. Fourteen percent of livers exhibited an increased share of CYP3A5 transcripts (range 4-20%). These livers were either heterozygous for the marker of the CYP3A5 polymorphism, the CYP3A5*1A allele, or expressed very low levels of CYP3A4 mRNA. The CYP3A7 expression was bimodal, and it was increased in 15% livers. CYP3A4 was the dominant CYP3A in the intestine, followed by CYP3A5. CYP3A5 and CYP3A7, but not CYP3A4, were also expressed in the adrenal gland and in the prostate, whereas only CYP3A5 was detected in the kidney. These three tissues were shown to express much lower levels of pregnane X receptor mRNA than the intestine, indicating possibly a different mode of regulation of CYP3A expression. Expression of CYP3A genes was undetectable in peripheral blood lymphocytes. In summary, these assays and results should aid in our efforts to further dissect the regulation and the physiological and pharmacological significance of CYP3A isozymes.

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“…In fact, the DexRE-1 of the rabbit and human CYP3A6 and CYP3A4, respectively, confers dexamethasone responsiveness on a heterologous promoter (Barwick et al, 1996;Lehmann et al, 1998). The DexRE-2 and Site A elements, however, are present only in mouse and rat isoforms (Burger et al, 1992;Telhada et al, 1992;Hashimoto et al, 1993;Jounaïdi et al, 1994;Barwick et al, 1996;Huss et al, 1996;Toide et al, 1997).…”

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Two-Stage Glucocorticoid Induction ofCYP3A23through Both the Glucocorticoid and Pregnane X Receptors

Huss

Kasper

2000

Mol Pharmacol

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Glucocorticoid inducibility of the CYP3A23 gene is conferred by a multisite unit comprising binding sites for several members of the nuclear receptor superfamily of transcription factors, including the chicken ovalbumin upstream promoter-transcription factor COUP-TF, pregnane X receptor (PXR), and hepatocyte nuclear factor 4 (HNF-4). The presence of three binding sites, each of which interacts with more than one factor, contributes to the complexity of the CYP3A23 glucocorticoid-responsive region. Despite the glucocorticoid sensitivity of this gene, direct binding of ligand-activated glucocorticoid receptor (GR) to the CYP3A23 dexamethasone-responsive region (DexRE) is not required for induction. This study demonstrates that DexRE-2 is the key element within the CYP3A23 proximal promoter, conferring ligand sensitivity via its interaction with the PXR/RXR␣ heterodimer. The DexRE-1 and HNF-4 sites are not ligand-responsive, but are essential accessory elements required for full promoter inducibility. In addition to ligandmediated activation of PXR, the overall induction response involves a GR-mediated stimulation of PXR and RXR␣ expression. Hence, the induction pathway can be divided into two stages. In stage one, maximal induction requires a GR-dependent increase in PXR and RXR␣ expression, and stage two is characterized by direct transcriptional activation of CYP3A23, which is dependent on ligand-activated PXR as well as accessory factors bound at the DexRE-1 and HNF-4 sites. Because multiple proteins bind at each element within the glucocorticoid-responsive region, factors not contributing to ligand responsiveness, such as chicken ovalbumin upstream promotertranscription factor, may modulate the response through competitive interactions.

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Effect of dexamethasone and phenobarbital on run-on transcription rate and CYP3A mRNA concentration in rat liver: Changes during development

Cited by 43 publications

References 40 publications

Nuclear Receptor Involvement in the Regulation of Rat Cytochrome P450 3A23 Expression

Nuclear Receptor Involvement in the Regulation of Rat Cytochrome P450 3A23 Expression

Interindividual Variability and Tissue-Specificity in the Expression of Cytochrome P450 3A mRNA

Two-Stage Glucocorticoid Induction ofCYP3A23through Both the Glucocorticoid and Pregnane X Receptors

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