Multiplicity of deoxyribonucleic acid sequences with homology to a cloned complementary deoxyribonucleic acid coding for rat phenobarbital-inducible cytochrome P-450

Mizukami, Yuzuru; Fujii‐Kuriyama, Yoshiaki; Muramatsu, Masami

doi:10.1021/bi00274a036

Cited by 63 publications

(32 citation statements)

References 13 publications

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“…3 (1984) genes for other forms of cytochrome P-450 or divergent P450Oc genes, and further studies to identify and characterize clones in a bovine genomic library should clarify this point. In the case of the phenobarbital subset of rat liver cytochromes P-450, there appear to be six to eight genes or pseudogenes encoding several forms of cytochrome P-450 (10)(11)(12). The identification and characterization of cDNA clones specific for P-450sc provide the tools necessary to examine the mechanisms by which trophic hormones regulate the synthesis of this form of cytochrome P-450.…”

Section: Resultsmentioning

confidence: 99%

“…Elucidation of the structure and organization of genes encoding certain xenobiotic-inducible forms of cytochrome P-450 has brought to light several interesting features including the existence of multiple genes (10)(11)(12) and unusually large mRNA molecules (13-15) as well as possible mechanisms involved in genetic regulation (11,12,(16)(17)(18)(19). However, little is known concerning the structure and organization of the genes and.…”

Section: Introductionmentioning

confidence: 99%

“…Linearized plasmid DNA (5 A.g/ 10 ,ul of H20) was denatured by boiling for 10 min and then was rapidly cooled on ice. The solution was made 0.5 M in NaOH and kept at room temperature for 20 min, neutralized by addition of sodium acetate, and immediately frozen on dry ice.…”

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confidence: 99%

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Identification and characterization of cDNA clones specific for cholesterol side-chain cleavage cytochrome P-450.

John¹,

Ashley²,

MacDonald³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

109

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Two overlapping cDNA clones (pBSCC-1 and pBSCC-2) bearing inserts -425 and ':950 base pairs long, respectively, which are specific for bovine cholesterol sidechain cleavage cytochrome P-450 (P-450CC), have been identified by using two differential hybridization screening procedures followed by hybrid-selected RNA translation. By using these cloned cDNAs as hybridization probes, an RNA species was identified that had the properties expected of mRNA specific for P-450, with respect to tissue specificity, corticotropin (ACTH)-mediated regulation of synthesis, and size of the protein product synthesized in vitro. In RNA samples obtained from bovine adrenal cortex, from bovine corpus luteum, and from cultured bovine adrenocortical cells, it was found that P450s is encoded by mRNA species -2000 bases long, a mujority of which are polyadenylylated. P-450s, mRNA was not detected in RNA samples prepared from bovine heart, liver, and kidney. Treatment of cultured bovine adrenocortical cells with ACTH resulted in the appearance of elevated levels of P-450, mRNA within 8 hr. Thus, ACTH promotes the enhancement of P-450scc gene transcription or acts to stabilize the transcripts. When pBSCC-2 cDNA was used to probe high molecular weight bovine DNA following treatment with restriction endonucleases, a simple pattern of hybridization was observed indicating that P-450O may be encoded by a single gene.Cytochromes P-450 are a diverse group of heme-containing mixed-function oxidases that are involved in the biotransformation of a wide variety of exogenous and endogenous substrates. The best-studied examples of those forms that metabolize endogenous substrates are the group of cytochromes P-450 required for steroid hormone biosynthesis. In steroidogenic tissues such as adrenal cortex, testis, ovary, and placenta, the initial and rate-limiting step in the pathway leading from cholesterol to steroid hormones is the cleavage of the side chain of cholesterol to yield pregnenolone (1). This reaction, known as cholesterol side-chain cleavage, is catalyzed by a specific form of cytochrome P-450 (P-450,cc) that is localized in the inner mitochondrial membrane (2,3). This protein is synthesized on cytoplasmic ribosomes as a higher molecular weight precursor (Mr 54,500) that is proteolytically processed to the mature form (Mr 49,000) upon insertion into the mitochondrion (4,5). By utilizing bovine adrenocortical cells in primary monolayer culture, it has been demonstrated in this laboratory that the synthesis of P-450,"c can be modulated by corticotropin (ACTH) and that this action of ACTH is mediated by cyclic AMP (6,7). A similar pattern of regulation of P-450Scc synthesis has been observed upon stimulation of bovine ovarian granulosa cells with follicle-stimulating hormone (8) and rat testicular Leydig cells with human chorionic gonadotropin (9). Thus, it appears that the levels of P-4Sscc are maintained in vivo in response to trophic hormone stimulation and that cyclic AMP plays a key role in the regulation of P-45scc synth...

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Identification and characterization of cDNA clones specific for cholesterol side-chain cleavage cytochrome P-450.

John¹,

Ashley²,

MacDonald³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

109

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“…Nearly half of these differences are contained within two short regions of relatively high (23-24%) sequence divergence that account for nearly half of the amino acid differences between the two polypeptides and are located within exon 7 ofthe cytochrome P-450e gene (17,18). These flank a more highly conserved segment encoding an "analogous tridecapeptide" observed by Ozols et al (19) in two ostensibly unrelated rabbit liver P450 isozymes, one constitutive (LM-3b) and one highly induced by phenobarbital .Recently, a series of genes distinct from the P-450e and P-450b genes but having sequence homology to cytochromes P-450b and P-450e cDNAs have been cloned and characterized (18,20). Hybridization of several of these genes to P-450e cDNA probes combined with heteroduplex analysis revealed that the region of high homology to the cytochrome P-450e gene is limited to exons 7 and 8, and possibly the intron between them (18).…”

mentioning

confidence: 94%

“…Recently, a series of genes distinct from the P-450e and P-450b genes but having sequence homology to cytochromes P-450b and P-450e cDNAs have been cloned and characterized (18,20). Hybridization of several of these genes to P-450e cDNA probes combined with heteroduplex analysis revealed that the region of high homology to the cytochrome P-450e gene is limited to exons 7 and 8, and possibly the intron between them (18).…”

Section: Introductionmentioning

confidence: 99%

Gene conversion in a cytochrome P-450 gene family.

Atchison¹,

Adesnik²

1986

Proc. Natl. Acad. Sci. U.S.A.

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The mRNAs encoding the two major phenobarbital-inducible forms of cytochrome P-450 of rat liver, P-450b and P-450e, are remarkably similar (98% homologous) in nucleotide sequence, but the distribution of differences within them is not random. While the 5' halves (s1 kilobase) appear to be identical, there are 36 divergent residues in the remaining sequences of the two mRNAs, with 14 differences residing in two short highly divergent segments, which in the P-450e gene are located within exon 7. DNA sequence analysis of portions of a number of P-450b/e-related genes provides strong evidence that at least one ofthe short divergent segments is the result of a recent gene conversion event between an ancestor to the cytochrome P-450e gene and a related donor P450 gene of unknown function. The sequence data also suggest that extensive gene conversion has occurred within all the members of this gene family in the region including exons 7 and 8 and the intron between them, with a resultant homogenization of those sequences relative to other portions of the genes. Genomic Southern blotting analysis demonstrates that the presence of an apparent "constant" region in the 5' halves of the P-450b and P-450e mRNAs does not reflect a rearrangement in somatic cells of a germ-line DNA configuration. It is therefore proposed that it, too, is a consequence of a very recent gene conversion event between ancestors of the genes encoding both proteins or of an unequal crossing-over between them. On the basis of these and other data we propose that gene conversion represents an important evolutionary mechanism for the generation of related cytochrome P-450 isozymes in which regions of extraordinary sequence similarity and dissimilarity are intermixed. The gene conversion mechanism would account for some of the overlaps in substrate specificities of distantly related P-450s as well as for substantial differences in catalytic properties between closely related members of the same P450 protein family.It has long been recognized (1) that members of a multigene family within a single species are generally much more closely related to each other than to equivalent (orthologous) genes in other species. This concerted evolution ofgenes that have had at least as long a time to diverge from each other as from their orthologous counterparts may result from unequal crossing-over between tandemly linked genes or from either double reciprocal recombination or gene conversion (1). The latter term refers to a nonreciprocal recombination event in which a segment of one gene replaces the corresponding segment of a related gene (2). The occurrence of homogenizing gene conversion events in the human a-globin (3), human fetal '-tglobin (4), oxytocin-vasopressin (5), and immunoglobulin (6) gene families has been inferred from a comparison of DNA sequences.t It has recently become apparent that gene conversion may also produce short regions of high diversity within closely related members of a multigene family, when the donor gene for the conversion is...

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Metabolism-Dependent Toxicities of Cyclophosphamide and Protection by N-Acetylcysteine and Other Thiols

Gurtoo¹,

Berrigan²,

Love³

et al. 1985

Experimental and Clinical Progress in Cancer Chemotherapy

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Multiplicity of deoxyribonucleic acid sequences with homology to a cloned complementary deoxyribonucleic acid coding for rat phenobarbital-inducible cytochrome P-450

Cited by 63 publications

References 13 publications

Identification and characterization of cDNA clones specific for cholesterol side-chain cleavage cytochrome P-450.

Identification and characterization of cDNA clones specific for cholesterol side-chain cleavage cytochrome P-450.

Gene conversion in a cytochrome P-450 gene family.

Metabolism-Dependent Toxicities of Cyclophosphamide and Protection by N-Acetylcysteine and Other Thiols

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