PubMed Health⌕ Search

Biomedical subjects

D W Nebert

Publications and source records attributed to D W Nebert.

At least 181 records · Page 10Linked to original sources
PubMed1985

Assignment of dioxin-inducible cytochrome P-450 gene family to Chinese hamster chromosome 4.

The 2,3,7,8-tetrachlorodibenzo-p-dioxin-inducible cytochrome P-450 gene family (P1-450 and P3-450 in the C57BL/6N mouse) has recently been localized to mouse chromosome 9. In the present study, HindIII-digested DNA from Chinese hamster, mouse, and 20 Chinese hamster X mouse somatic cell hybrids and subclones segregating hamster chromosomes was probed with the mouse P1-450 and P3-450 full-length cDNA clones. Hamster P-450 gene fragments (6.0 and 7.4 kb) were assigned to Chinese hamster chromosome 4. These data are consistent with linkage conservation among these two P-450 sequences and four other loci on mouse chromosome 9 that map to hamster chromosome 4.

Animals↗
PubMed1985

Developmental and tissue-specific differential regulation of the mouse dioxin-inducible P1-450 and P3-450 genes.

The murine Ah locus has two structural genes, P1-450 and P3-450, that are members of the 2,3,7,8-tetrachlorodibenzo-p-dioxin-inducible P-450 gene family and are closely linked on mouse chromosomes 9. Transcriptional activation of both genes in liver is controlled by the Ah receptor. Inducible P1-450 mRNA is correlated with aryl hydrocarbon hydroxylase activity, whereas inducible P3-450 mRNA is correlated with acetanilide 4-hydroxylase activity. This report shows that inducible P3-450 mRNA is also highly correlated with estradiol 2-hydroxylase activity. Both P1-450 and P3-450 activities are induced by 3-methylcholanthrene in liver; in contrast, P1-450 but not P3-450 activity is induced to a significant extent in kidney, lung, and intestine. Constitutive levels of P3-450 mRNA in liver are at least five times greater than those of P1-450 mRNA, thus accounting for the much greater "fold inducibility" of aryl hydrocarbon hydroxylase than acetanilide 4-hydroxylase or estradiol 2-hydroxylase. The hepatic P3-450 induction response occurs at lower inducer concentrations than the P1-450 induction response, suggesting a difference in affinity of the inducer-receptor complex for regulatory regions of the two genes. Developmentally, P1-450 inducibility occurs at least 2 weeks earlier in gestation than P3-450 inducibility, which occurs near the time of birth. These data thus demonstrate striking differences between the expression of two homologous genes in the same P-450 subfamily, with respect to developmental and tissue specificity and sensitivity to common inducers that interact with the Ah receptor.

Animals↗
PubMed1985

Comparison of the mouse P(1)450 gene and flanking sequences from a MOPC 41 plasmacytoma and normal liver.

The murine P(1)450 gene is inducible by foreign chemicals such as dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin; TCDD). The induction process appears to involve binding of TCDD to the Ah receptor, interaction of the inducer-receptor complex with chromatin, and transcriptional activation of the P(1)450 gene. P(1)450 gene expression did not occur in the MOPC 41 plasmacytoma transplanted intra-abdominally in the BALB/c mouse. By Southern blot analysis, the P(1)450 gene restriction patterns are similar between MOPC 41 DNA and normal mouse liver DNA digested with Eco RI, Bam HI, or Hind III, suggesting there is no evidence for a gross chromosomal rearrangement in the tumor P(1)450 gene. The P(1)450 gene isolated from a genomic DNA library derived from the MOPC 41 tumor lacked the first exon and two-thirds of the first intron because of a rearrangement that presumably occurred during construction of the genomic DNA library. An unknown upstream segment of 2578 bases that had recombined with the remainder of the P(1)450 gene was sequenced and found to contain several repetitive sequences on both strands. The remaining 6272 bases of the P(1)450 gene (2532 bp of exons, 1969 bp of introns and 1771 bp of 3'-flanking region) are identical between the BALB/cJ-derived MOPC 41 and normal C57BL/6N mouse liver, with the exception of one CTTT repeat in the second intron. Hence, the BALB/c and C57BL/6 inbred mice, generally regarded as among the most divergent of Mus domesticus inbred laboratory strains, possess a P(1)450 gene and flanking region that exhibit greater than 99.9% similarity in nucleotide sequence.

Animals↗
PubMed1984

Regulation of mouse cytochrome P3-450 by the Ah receptor. Studies with a P3-450 cDNA clone.

The Ah locus in the C57BL/6N mouse regulates at least two cytochrome P-450 gene products, termed in the mouse P1-450 and P3-450; these two enzymes are so named because each is responsible for the highest turnover number for the substrates benzo[a]pyrene and acetanilide, respectively. A cDNA library was prepared in pBR322 from sucrose gradient fractionated total liver poly(A+)-enriched RNA (approximately 20 S) from 2,3,7,8-tetrachlorodibenzo-p-dioxin- (TCDD) treated C57BL/6N (Ahb/Ahb) mice. Differential colony hybridization screening, with [32P]cDNA probes derived from total liver mRNA of both TCDD-treated and control C57BL/6N mice, yielded pP(3)450-21 (1710 base pair) and pP(1)450-57 (1770 base pair) cDNA clones. pP(1)450-57 was found to have 690 base pairs 5'-ward of the original P1-450 cDNA cloned in this laboratory. Restriction maps of pP(3)450-21 and pP(1)450-57 are markedly different and clearly are derived from separate genes. By means of hybridization-translation-arrest experiments, anti-(P3-450) precipitates the translation product (Mr approximately equal to 55000) of mRNA specifically hybridizing to pP(3)450-21. It is also shown that hybridization-translation-arrest experiments using polyclonal antibodies are not specific for proof of a P-450 cDNA clone. pP(3)450-21 was used to probe liver mRNA from Ahb/Ahb, Ahb/Ahd, and Ahd/Ahd mice treated with 3-methylcholanthrene, beta-naphthoflavone, aroclor 1254, isosafrole, low TCDD, or high TCDD. These genetic data rigorously demonstrate control of the P3-450 (20S) mRNA induction process by the Ah receptor. pP(3)450-21 fragments hybridized to TCDD-induced C57BL/6N mRNA and to a portion of the cloned 5' end of the P1-450 gene from a mouse MOPC 41 plasmacytoma library.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Purification and characterization of a microsomal cytochrome P-450 with high activity of coumarin 7-hydroxylase from mouse liver.

Phenobarbital-induced coumarin 7-hydroxylase is high in DBA/2J and low in C57BL/6N inbred mice; this genetic difference is encoded by the Coh locus on chromosome 7. The aim of this study was to develop an antibody specific for this cytochrome P-450 polymorphism. P-450 fractions, highly specific for phenobarbital-inducible coumarin 7-hydroxylase activity, were purified from DBA/2J and C57BL/6N mouse liver microsomes. Both proteins are 49 kDa, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Soret peaks of the reduced cytochrome . CO complexes are 451 nm. Reconstituted DBA/2J coumarin 7-hydroxylase activity exhibits a V twice as high as, and a Km value 10-fold less than, the reconstituted C57BL/6N activity. Antibodies were raised in rabbit. By Ouchterlony immunodiffusion, both antibodies show 100% cross-reactivity with DBA/2J and C57BL/6N microsomes and purified antigens. Yet, DBA/2J but not C57BL/6N 7-hydroxylase activity is inhibited by the antibody to DBA/2J P-450. Both DBA/2J and C57BL/6N activities are blocked by the antibody to C57BL/6N P-450. Neither antibody has any effect on liver microsomal d-benzphetamine N-demethylase, ethylmorphine N-demethylase, aminopyrine N-demethylase, 7-ethoxycoumarin O-deethylase, acetanilide 4-hydroxylase, or aryl hydrocarbon (benzo[a]pyrene) hydroxylase activity. The DBA/2J protein most specific for phenobarbital-induced coumarin 7-hydroxylation is designated 'P-450Coh'. Anti-(P-450Coh) precipitates a relatively minor 49-kDa protein from detergent-solubilized microsomes and from in vitro translation of poly(A+)-enriched total RNA of phenobarbital-treated DBA/2J mouse liver, whereas the major phenobarbital-induced P-450 proteins exhibit a molecular mass of about 51 kDa. The immunoprecipitated translation products correspond to a messenger RNA of 2100 +/- 100 nucleotides.

Animals↗
PubMed1984

The murine Ah locus. Comparison of the complete cytochrome P1-450 and P3-450 cDNA nucleotide and amino acid sequences.

Mouse cytochrome P1-450 and P3-450 are most closely associated with induced aryl hydrocarbon (benzo[a]pyrene) hydroxylase (EC 1.14.14.1) and acetanilide 4-hydroxylase activity, respectively. Full-length cDNA clones of P1-450 and P3-450 were generated from mRNA isolated from 3-methylcholanthrene-treated C57BL/6N mouse liver. P1-450 cDNA is 2620 nucleotides in length and has a coding region (base 110 to 1,675) that produces a protein with 521 residues (Mr = 58,914). P3-450 cDNA is 1,894 nucleotides in length and yields a protein with 513 residues (Mr = 58,183). P1-450 mRNA is the first reported example in mouse in which UAG is used as the termination codon. P1-450 and P3-450, both induced by polycyclic hydrocarbons and regulated by the Ah receptor, exhibit overall nucleotide and protein homology of 68, and 73%, respectively. Segments of high homology, interspersed with regions of low homology, support the hypothesis of gene conversion or unequal crossing over as possible mechanisms for divergence of these two genes. Mouse P1-450 and P3-450 cDNAs were compared with previously published data on rat P-450e cDNA and rabbit form 2 protein, corresponding to two P-450 genes from the "phenobarbital inducible" P-450 gene subfamily. Nucleotide homology between a member of either gene subfamily is about 30%, and protein homology is about 15%, suggesting that the Ah locus-associated P-450 gene subfamily diverged from the phenobarbital inducible P-450 subfamily more than 200 million years ago. An N-terminal and a C-terminal cysteinyl fragment corresponding to the regions around P1-450 Cys-158 and Cys-458, respectively, are the only two cysteinyl peptides conserved among all four proteins compared. Because of greater homology in the C-terminal conserved cysteinyl fragment between the two gene subfamilies and a greater hydrophobic pocket in the C-terminal conserved cysteinyl fragment, the data favor this cysteine as the more likely candidate for the thiolate ligand to the heme iron in the P-450 enzyme active-site.

Amino Acid Sequence↗
PubMed1984

Changes in cytochrome P-450 iron spin state as a function of castration, testosterone or estrogen treatment, vigorous exercise, or starvation.

3-Methylcholanthrene treatment induced large increases in high-spin P-450 form(s) in intact microsomal membranes from New Zealand White rabbit or C57BL/6N mouse liver but not in rabbit lung, kidney or intestine. Testosterone enhanced the high-spin/low-spin ratio of P-450 in female C57BL/6N liver to the same degree as 3-methylcholanthrene treatment; there was no additive increase in the ratio with treatment of testosterone plus 3-methylcholanthrene. The same effect was seen in ovariectomized females. In the liver of normal or castrated C57BL/6N mice, estradiol-17 beta pretreatment caused no significant change in the high-spin/low-spin ratio. Vigorous homogenization of the microsomes can decrease the high-spin/low-spin ratio, more so in control than in 3-methylcholanthrene-induced C57BL/6N microsomes. Vigorous exercise (20-min swim) and fasting for 2 days significantly enhanced the high-spin/low-spin ratio in C57BL/6N liver. Temperature studies showed that the low-spin electron paramagnetic resonance signal intensity, as a function of absolute temperature in the range of 16 to 70 degrees K, was consistent with a simple Boltzmann dependence within the Kramers doublet giving rise to the microwave transition. We conclude that, within the limits of measurement, the low-spin state acts like an isolated state and is not in thermal equilibrium with the high-spin form of P-450.

Animals↗
PubMed1984

Mouse cytochrome P3-450: complete cDNA and amino acid sequence.

A full-length cDNA clone (1,894 nucleotides) of mouse cytochrome P3-450 was isolated with the Okayama-Berg vector and sequenced. An open reading frame spanned positions 61 to 1602. The first 25, and three of the last five, amino acids of P3-450 are identical to those found in the amino- and carboxy-terminus, respectively, of the rat P-450d protein. Mouse P3-450 protein has 513 residues, and a molecular weight of 58,223 with six cysteine residues. P3-450 nucleotides 305 to 352 exhibit 74% homology, and nucleotides 1068 to 1260, 69% homology, with portions of rat P-450b exons 2 and 7, respectively. P3-450 shows 62% homology in the so-called "highly conserved region" of 39 nucleotides in the rat P-450b and P-450e and the mouse P-450b. These results indicate that P3-450, P-450b and P-450e arose from a common ancestral gene. Cysteinyl peptide-coding regions were examined: P3-450 nucleotides 1405 to 1464 exhibit 61% homology, and nucleotides 502 to 552 exhibit 37% homology, when compared with their corresponding regions in the rat P-450b gene. These data support the likelihood that cysteine 456 is the thiolate ligand to the heme iron in the P3-450 enzyme active-site.

Amino Acid Sequence↗
PubMed1984

The Ah receptor: binding specificity only for foreign chemicals?

The murine Ah locus controls the induction of at least four drug-metabolizing enzymes: cytochromes P1-450, P2-450, and P3-450, and UDP-glucuronosyltransferase. The Ah gene codes for a cytosolic receptor. It is known that the induction response includes: (i) high-affinity binding of specific foreign chemicals to the Ah receptor; (ii) temperature-dependent translocation of the "activated" inducer-receptor complex into the nucleus; (iii) binding of the complex presumably to chromatin components; (iv) transcriptional activation of specific genes; (v) maximal increases in intranuclear high-molecular-weight precursor mRNA (pre-mRNA) that precede by several hours the maximal increases in cytoplasmic mRNA; (vi) translation of the mRNA principally on membrane-bound polysomes; and (vii) increases in the specific membrane-bound proteins (including architectural arrangement with other membrane-bound moieties) that reflect enhanced specific drug-metabolizing activities. It is not known how many of the other drug metabolism induction responses are also governed by receptors. The Ah locus studies have been chiefly unraveled in the mouse, due to several inbred strains having a receptor defect. In addition to "classical" pharmacologic methods (such as structure-activity studies) and standard biochemical techniques, the newer methods of recombinant DNA technology and somatic-cell genetics in culture are shown to be important in understanding the Ah receptor and its induction response. It is possible that this receptor is required for endogenous functions critical to life processes, as well as its function in the induction of drug metabolism by certain polycyclic aromatic compounds.

Animals↗
PubMed1984

Isosafrole-induced cytochrome P2-450 in DBA/2N mouse liver. Characterization and genetic control of induction.

Mouse "cytochrome P2-450" is defined as that form of isosafrole-induced P-450 in DBA/2N liver most specifically correlated with isosafrole metabolism. Isosafrole pretreatment does not induce aryl hydrocarbon hydroxylase activity ("cytochrome P1-450") in C57BL/6N or DBA/2N mice, induces acetanilide 4-hydroxylase activity ("cytochrome P3-450") more than 3-fold in C57BL/6N but not in DBA/2N mice, and induces isosafrole metabolite formation more than 3-fold in both C57BL/6N and DBA/2N mice. P2-450 was, therefore, purified from isosafrole-treated DBA/2N liver microsomes having negligible amounts of contaminating P1-450 and P3-450. The apparent molecular weight of P2-450 is 55,000, and the protein appears homogeneous on sodium dodecyl sulfate-polyacrylamide gels. The Soret peak of the reduced purified cytochrome X CO complex is 448 nm. Purified P2-450, reconstituted in vitro, metabolizes acetanilide poorly and benzo[a]pyrene hardly at all. Anti-(P2-450) inhibits (90 to 100%) liver microsomal isosafrole metabolite formation, yet has no effect on aryl hydrocarbon hydroxylase, acetanilide 4-hydroxylase, biphenyl 2- or 4-hydroxylase, or 7-ethoxycoumarin O-de-ethylase activities. 3-Methylcholanthrene induces anti-(P2-450)-precipitable protein about 12-fold in C57BL/6N and 2-fold in DBA/2N liver; 2,3,7,8-tetrachlorodibenzo-p-dioxin (10 micrograms/kg), about 12-fold in both C57BL/6N and DBA/2N liver; isosafrole, more than 3-fold in both C57BL/6N and DBA/2N. Benzo[a]anthracene at maximal doses induces anti-(P2-450)-precipitable protein in C57BL/6N liver no more than 2-fold, yet is known to be a highly potent inducer of P1-450 mRNA in C57BL/6N liver. The sensitivity of the P2-450 induction process to isosafrole is inherited as an autosomal additive trait; studies of offspring from the C57BL/6N(DBA/N)F1 X DBA/2N backcross confirm involvement of the Ah locus or s closely segregating gene. In contrast, among crosses between C57BL/6N and DBA/2N, sensitivity of the P1-450 and P3-450 induction process to 3-methylcholanthrene or 2,3,7,8-tetrachlorodibenzo-p-dioxin is inherited as an autosomal dominant trait. These data suggest that, although P1-450, P2-450, and P3-450 proteins are controlled by the Ah locus, either a P-450 protein polymorphism exists between C57BL/6N and DBA/2N mice or subtle differences may exist in the interaction of various inducers with Ah receptor.

Animals↗
PubMed1984

Importance of the route of administration for genetic differences in benzo[a]pyrene-induced in utero toxicity and teratogenicity.

C57BL/6N (Ahb/Ahb) mice have a high-affinity Ah receptor in tissues, whereas AKR/J and DBA/2N (Ahd/Ahd) mice have a poor-affinity Ah receptor. The cytochrome P1-450 induction response (enhanced benzo[a]pyrene metabolism) occurs much more readily in Ahb/Ahb and Ahb/Ahd than in Ahd/Ahd mice, at any given dose of the inducer benzo[a]pyrene. Embryos from the AKR/J X (C57BL/6N)(AKR/J)F1 and the reciprocal backcross were studied during benzo[a]pyrene feeding of the pregnant females. Oral benzo[a]pyrene (120 mg/kg/day) given to pregnant Ahd/Ahd mice between gestational day 2 and 10 produces more intrauterine toxicity and malformations in Ahd/Ahd than Ahb/Ahd embryos. This striking allelic difference is not seen in pregnant Ahb/Ahd mice receiving oral benzo[a]pyrene. Pharmacokinetics studies with [3H]benzo[a]pyrene in the diet and high-performance liquid chromatographic analysis of benzo[a]pyrene metabolism in vitro by the maternal intestine, liver, and ovary and the embryos of control and oral benzo[a]pyrene-treated pregnant females are consistent with "first-pass elimination" kinetics and differences in benzo[a]pyrene metabolism by the embryos and/or placentas versus maternal tissues. In the pregnant Ahd/Ahd mouse receiving oral benzo[a]pyrene, little induction of benzo[a]pyrene metabolism occurs in her intestine and liver; this leads to much larger amounts of benzo[a]pyrene reaching her embryos, and genetic differences in toxicity and teratogenesis are manifest. In the pregnant Ahb/Ahd mouse receiving oral benzo[a]pyrene, benzo[a]pyrene metabolism is greatly enhanced in her intestine and liver; this leads to less benzo[a]pyrene reaching her embryos, much less intrauterine toxicity and malformations, and no genetic differences are manifest. More toxic metabolites (especially benzo[a]pyrene 1,6- and 3,6-quinones) are shown to occur in Ahd/Ahd embryos than in Ahb/Ahd embryos. In additional studies, no prenatal or neonatal "imprinting" effect in C57BL/6N mice by 2,3,7,8-tetrachlorodibenzo-p-dioxin or Aroclor 1254 on benzo[a]pyrene metabolism later in life was detectable. These genetic differences in intrauterine toxicity and teratogenicity induced by oral benzo[a]pyrene are just opposite those induced by intraperitoneal benzo[a]pyrene [Shum et al., '79; Hoshino et al., '81). The data in the present report emphasize the importance of the route of administration when the teratogen induces its own metabolism.

Abnormalities, Drug-Induced↗
PubMed1984

Induction of immunotoxicity by polycyclic hydrocarbons: role of the Ah locus.

We have employed the plaque forming cell (PFC) response to sheep erythrocytes as well as lymphocyte proliferation to study the induction of immunotoxicity in AHH-inducible (Ah Locus positive, C57BL/6N; B6C3F1) and AHH non-inducible (Ah Locus negative, DBA2/N) mice following administration of polycyclic aromatic hydrocarbons. When two potent carcinogenic polycyclic hydrocarbons which induce AHH activity, 3-methylcholanthrene (MCA) or 1,2,5,6-dibenzanthracene [DB(a,h)A] were administered IP, immunotoxicity was observed in both AHH-inducible and AHH non-inducible animals. However, the AHH-inducible animals appeared to be more sensitive, and substantial suppression of a PFC response toxicity could be induced with doses as low as 14 mg/kg methylcholanthrene. While suppression of a mitogen response required a dose of 43-125 mg/kg. Administration of the weak carcinogen 1,2,3,4-dibenzanthracene [DB(a,c)A], IP, which similarly induces AHH activity in inducible animals, failed to induce immunotoxicity in either C57B1/6N or DBA/2N mice. In contrast to the results obtained following IP administration, when MCA was administered repeatedly (4X) via an intragastric (IG) route we observed striking immunosuppression of a PFC response in Ah locus negative (DBA/2) animals but minimal effects in Ah locus positive animals (C57B1/6). We finally observed that a single IP dose of MCA (125 mg/kg) to Ah locus positive animals substantially inhibited Natural Killer Cell activity but had more limited effects on the ability of an animal to reject a challenge by an immunogenic syngeneic fibrosarcoma.

Animals↗
PubMed1984

Isolation and characterization of full-length mouse cDNA and genomic clones of 3-methylcholanthrene-inducible cytochrome P1-450 and P3-450.

Polysome immuno-adsorption, with immunoglobulin G directed against two 3-methylcholanthrene-induced mouse liver cytochrome P-450 proteins, was used to enrich mRNA from 3-methylcholanthrene-treated C57BL/6N mouse liver. cDNA transcribed from the P-450-enriched mRNA was then cloned into the Okayama-Berg vector. Two cDNA classes were detected upon differential screening of the clone bank with [32P]cDNA derived from 3-methylcholanthrene-induced immuno-enriched versus control mRNA. Several representatives of these two classes were judged to be near full length by comparison with their corresponding mRNA mobilities on denaturing agarose gels. A continuous reading-frame near the 5' end of one cDNA class (P1-450) corresponds to a protein having 15 of 17 residues the same as the published N-terminal sequence of rat P-450c. A continuous reading frame near the 5' end of the other class (P3-450) corresponds exactly to the first 25 amino acids of the published N-terminal sequence of rat P-450d. The P1-450 cDNA is at least 700 bp longer than the P3-450 cDNA. Heteroduplex analysis and Southern blot hybridization demonstrate that these mRNAs share approx. 1100 bp of sequence homology. Genomic P1-450 and P3-450 clones were isolated from a gene library constructed from C57BL/6N mouse liver DNA. By heteroduplex analysis with the corresponding cDNA, the P1-450 gene spans about 6 kb and the P3-450 gene about 7 kb. The intron-exon patterns are very similar, with the second and seventh exons being much larger than the other five. The 3' terminal exon of P1-450 is about 500 bp longer than that of P3-450. These data suggest that both P1-450 and P3-450 have diverged from a common ancestral gene.

Animals↗
PubMed1984

Localization of cytochrome P1-450 and P3-450 genes to mouse chromosome 9.

Treatment of mice with polycyclic aromatic hydrocarbons results in the induction of P1-450 and P3-450 forms of cytochrome P-450. The genes for both cytochromes have recently been cloned and shown to be coordinately regulated by the Ah receptor. The mouse analogues of P1-450 and P3-450 can be distinguished from their hamster counterparts by Southern blot analysis with Kpn I-digested DNA fragments. DNA from hamster-mouse somatic cell hybrids that have selectively lost mouse chromosomes was used in Southern blots to map the location of the two mouse genes. Chromosome segregation analysis of 12 hybrid clones demonstrated that the structural genes for both P1-450 and P3-450 can be assigned to mouse chromosome 9.

Animals↗
PubMed1984

Characterization of a cDNA clone for mouse phenobarbital-inducible cytochrome P-450b.

p40, a mouse 1040-nucleotide cDNA clone encoding a form of phenobarbital-inducible cytochrome P-450b, was selected by probing a cDNA library derived from phenobarbital-treated DBA/2N mouse liver with a rat 1830-nucleotide cDNA probe complementary to P-450b mRNA. When rat and mouse liver poly(A)+-enriched total RNAs are probed with p40, control rat P-450b mRNA levels are at least three times as much as control mouse P-450b levels. A 15-fold increase in rat 19S mRNA and only a 3-fold increase in the corresponding mouse 19S mRNA are found following phenobarbital treatment. An intranuclear 4800-nucleotide mRNA precursor is also detectable during the induction process with phenobarbital. Clofibrate, a hypolipidemic drug, is a potent inducer of P-450. There is no cross-hybridization between clofibrate-induced mRNA and either mouse P-450b cDNA or mouse P1-450 cDNA clones. At the level of hybridization stringency used for the Northern analysis, there is no cross-hybridization between phenobarbital-induced mouse or rat P-450b and the P1-450 cDNA probe or between 3-methylcholanthrene-induced mouse or rat P1-450 and the P-450b cDNA probe. These data indicate that the induction process by clofibrate involves activation of P-450 gene(s) not present in the multigene families inducible by either phenobarbital or polycyclic aromatic compounds, i.e., at least three sets of P-450 genes operate independently of one another. Two previously reported cDNA clones of phenobarbital-inducible rat P-450, R17 (P- 450e ) and pcP- 450pb4 (P-450b), were compared with mouse p40.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1984

Aryl hydrocarbon hydroxylase induction by benzo[a]anthracene: regulatory gene localized to the distal portion of mouse chromosome 17.

Aryl hydrocarbon (benzo[a]pyrene) hydroxylase inducibility by benzo[a]anthracene was studied in 29 somatic cell hybrid clones, developed by fusing mouse spleen or peritoneal cells from four different inbred strains with hypoxanthine phosphoribosyltransferase-deficient Chinese hamster E36 cells. Karyotype analysis plus 25 markers assigned to 16 autosomes and the X chromosome were examined. In 28 of the 29 clones, the presence or absence of inducibility is associated with the presence or absence, respectively, of mouse chromosome 17. Liver microsomal aryl hydrocarbon hydroxylase induction by 3-methylcholanthrene or benzo[a]anthracene was assessed in appropriate backcrosses with the Mus musculus molossinus, M. m. castaneus, MOR/Cv, PL/J, SM/J and DBA/2J inbred strains and in 13 NX8 recombinant inbred lines. Twenty-seven biochemical genetic markers representing all but four autosomes were tested for possible linkage with the hydroxylase inducibility, and no linkage was found. The hepatic Ah receptor was quantitated in 26 BXD recombinant inbred lines; the Ah phenotype did not match exactly any of the more than 70 genes with established strain distribution patterns representing 12 autosomes and at least five unlinked markers. It is concluded that a major gene controlling aryl hydrocarbon hydroxylase inducibility by benzo[a]anthracene is located on chromosome 17. Because there is no significant linkage with any of three biochemical markers in the upper third of the chromosome, we conclude that the inducibility gene is located in the distal 40% of mouse chromosome 17. Whether this trait represents the Ah locus, i.e., the gene encoding the cytosolic Ah receptor, will require further study.

Animals↗
PubMed1984

Association of fertility, fitness and longevity with the murine Ah locus among (C57BL/6N) (C3H/HeN) recombinant inbred lines.

The Ah locus encodes a cytosolic receptor which controls the induction of enzymes that metabolize drugs, chemical carcinogens, and other environmental pollutants. B6NXC3N recombinant inbred lines have been developed from the progenitors C57BL/6N and C3H/HeN inbred mouse strains. Ah phenotyping at each generation has resulted in the establishment of some lines containing high levels of the high-affinity Ah receptor; other lines, very low levels. A genetic model involving two unlinked loci is offered to explain the distribution of Ah receptor levels among (C57BL/6N) (C3H/HeN)F2 individuals. Between generations 7 and 13, individual females and males from the B6NXC3N recombinant inbred lines were crossed with DBA/2N males and females. Presence of high levels of the high-affinity Ah receptor in both female and male B6NXC3N mice was found to be associated with greater fertility, fitness, and longer life span. The data suggest that these parameters are correlated with the Ah locus or a closely segregating gene.

Animals↗