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. 1989 Apr 25;17(8):2907–2917. doi: 10.1093/nar/17.8.2907

Close linkage of the human cytochrome P450IIA and P450IIB gene subfamilies: implications for the assignment of substrate specificity.

J S Miles 1, W Bickmore 1, J D Brook 1, A W McLaren 1, R Meehan 1, C R Wolf 1
PMCID: PMC317701  PMID: 2726448

Abstract

We have isolated from human liver libraries two cytochrome P450 cDNA clones (lambda MP14 and lambda MP3) which are highly similar (83% over the coding region) to mouse testosterone 15 alpha hydroxylase and are therefore part of the cytochrome P450IIA gene subfamily. The P450IIA (CYP2A) gene subfamily was found to be closely linked to the P450IIB (CYP2B) subfamily and their chromosomal location could not be distinguished using somatic cell hybrids containing fragments of chromosome 19 between 19q12 and 19q13.2. Pulsed field gel electrophoresis indicates that both gene subfamilies are contained within 350-kb genomic DNA fragments, but were separated using various restriction enzymes. Northern blot analysis identified three P450IIA mRNAs each showing a wide inter-individual variation in their levels in the liver. High levels of P450IIA transcript were associated with high levels of P450IIB transcript suggesting that common factors may influence the expression of genes within these subfamilies. Genetic analysis has suggested previously that a member of the P450IIB subfamily is responsible for coumarin hydroxylase activity in the mouse. We discuss the possibility, based on our findings of tight linkage of the human P450IIA and IIB subfamilies, that a member of the IIA subfamily is a better candidate for this enzyme activity.

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Selected References

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  1. Adesnik M., Atchison M. Genes for cytochrome P-450 and their regulation. CRC Crit Rev Biochem. 1986;19(3):247–305. doi: 10.3109/10409238609084657. [DOI] [PubMed] [Google Scholar]
  2. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  3. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Brook J. D., Skinner M., Roberts S. H., Rettig W. J., Almond J. W., Shaw D. J. Further mapping of markers around the centromere of human chromosome 19. Genomics. 1987 Dec;1(4):320–328. doi: 10.1016/0888-7543(87)90031-0. [DOI] [PubMed] [Google Scholar]
  5. Carle G. F., Olson M. V. An electrophoretic karyotype for yeast. Proc Natl Acad Sci U S A. 1985 Jun;82(11):3756–3760. doi: 10.1073/pnas.82.11.3756. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Davis M. B., Schonk D., Monteiro M., Oerlemans F., Povey S., Wieringa B. Localization of PEPD to the long arm of chromosome 19. Ann Hum Genet. 1987 Jul;51(Pt 3):195–199. doi: 10.1111/j.1469-1809.1987.tb00871.x. [DOI] [PubMed] [Google Scholar]
  7. Davis M. B., West L. F., Shephard E. A., Phillips I. R. Regional localization of a human cytochrome P-450 (CYP1) to chromosome 19q13.1-13.3. Ann Hum Genet. 1986 Jul;50(Pt 3):237–240. doi: 10.1111/j.1469-1809.1986.tb01044.x. [DOI] [PubMed] [Google Scholar]
  8. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  9. Hill R. E., Shaw P. H., Barth R. K., Hastie N. D. A genetic locus closely linked to a protease inhibitor gene complex controls the level of multiple RNA transcripts. Mol Cell Biol. 1985 Aug;5(8):2114–2122. doi: 10.1128/mcb.5.8.2114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Juvonen R. O., Shkumatov V. M., Lang M. A. Purification and characterization of a liver microsomal cytochrome P-450 isoenzyme with a high affinity and metabolic capacity for coumarin from pyrazole-treated D2 mice. Eur J Biochem. 1988 Jan 15;171(1-2):205–211. doi: 10.1111/j.1432-1033.1988.tb13777.x. [DOI] [PubMed] [Google Scholar]
  11. Kumar A., Raphael C., Adesnik M. Cloned cytochrome P-450 cDNA. Nucleotide sequence and homology to multiple phenobarbital-induced mRNA species. J Biol Chem. 1983 Sep 25;258(18):11280–11284. [PubMed] [Google Scholar]
  12. Kwok S. C., Ledley F. D., DiLella A. G., Robson K. J., Woo S. L. Nucleotide sequence of a full-length complementary DNA clone and amino acid sequence of human phenylalanine hydroxylase. Biochemistry. 1985 Jan 29;24(3):556–561. doi: 10.1021/bi00324a002. [DOI] [PubMed] [Google Scholar]
  13. Leader D. P., Gall I., Lehrach H. The structure of a cloned mouse gamma-actin processed pseudogene. Gene. 1985;36(3):369–374. doi: 10.1016/0378-1119(85)90193-3. [DOI] [PubMed] [Google Scholar]
  14. Lindsay S., Bird A. P. Use of restriction enzymes to detect potential gene sequences in mammalian DNA. 1987 May 28-Jun 3Nature. 327(6120):336–338. doi: 10.1038/327336a0. [DOI] [PubMed] [Google Scholar]
  15. Matsunaga T., Nagata K., Holsztynska E. J., Lapenson D. P., Smith A., Kato R., Gelboin H. V., Waxman D. J., Gonzalez F. J. Gene conversion and differential regulation in the rat P-450 IIA gene subfamily. Purification, catalytic activity, cDNA and deduced amino acid sequence, and regulation of an adult male-specific hepatic testosterone 15 alpha-hydroxylase. J Biol Chem. 1988 Dec 5;263(34):17995–18002. [PubMed] [Google Scholar]
  16. Meehan R. R., Forrester L. M., Stevenson K., Hastie N. D., Buchmann A., Kunz H. W., Wolf C. R. Regulation of phenobarbital-inducible cytochrome P-450s in rat and mouse liver following dexamethasone administration and hypophysectomy. Biochem J. 1988 Sep 15;254(3):789–797. doi: 10.1042/bj2540789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Miles J. S., Spurr N. K., Gough A. C., Jowett T., McLaren A. W., Brook J. D., Wolf C. R. A novel human cytochrome P450 gene (P450IIB): chromosomal localization and evidence for alternative splicing. Nucleic Acids Res. 1988 Jul 11;16(13):5783–5795. doi: 10.1093/nar/16.13.5783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Nagata K., Matsunaga T., Gillette J., Gelboin H. V., Gonzalez F. J. Rat testosterone 7 alpha-hydroxylase. Isolation, sequence, and expression of cDNA and its developmental regulation and induction by 3-methylcholanthrene. J Biol Chem. 1987 Feb 25;262(6):2787–2793. [PubMed] [Google Scholar]
  19. Nebert D. W., Adesnik M., Coon M. J., Estabrook R. W., Gonzalez F. J., Guengerich F. P., Gunsalus I. C., Johnson E. F., Kemper B., Levin W. The P450 gene superfamily: recommended nomenclature. DNA. 1987 Feb;6(1):1–11. doi: 10.1089/dna.1987.6.1. [DOI] [PubMed] [Google Scholar]
  20. Nebert D. W., Gonzalez F. J. P450 genes: structure, evolution, and regulation. Annu Rev Biochem. 1987;56:945–993. doi: 10.1146/annurev.bi.56.070187.004501. [DOI] [PubMed] [Google Scholar]
  21. Nebert D. W., Nelson D. R., Adesnik M., Coon M. J., Estabrook R. W., Gonzalez F. J., Guengerich F. P., Gunsalus I. C., Johnson E. F., Kemper B. The P450 superfamily: updated listing of all genes and recommended nomenclature for the chromosomal loci. DNA. 1989 Jan-Feb;8(1):1–13. doi: 10.1089/dna.1.1989.8.1. [DOI] [PubMed] [Google Scholar]
  22. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  23. Phillips I. R., Shephard E. A., Ashworth A., Rabin B. R. Isolation and sequence of a human cytochrome P-450 cDNA clone. Proc Natl Acad Sci U S A. 1985 Feb;82(4):983–987. doi: 10.1073/pnas.82.4.983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rampersaud A., Walz F. G., Jr Mapping of genes for cytochromes P-450b, P-450e, P-450g, and P-450h in the rat. Demonstration of two gene clusters with P450-b,e localized on chromosome 1. J Biol Chem. 1987 Apr 25;262(12):5649–5653. [PubMed] [Google Scholar]
  25. Raunio H., Kojo A., Juvonen R., Honkakoski P., Järvinen P., Lang M. A., Vähäkangas K., Gelboin H. V., Park S. S., Pelkonen O. Mouse hepatic cytochrome P-450 isozyme induction by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene, pyrazole, and phenobarbital. Biochem Pharmacol. 1988 Nov 1;37(21):4141–4147. doi: 10.1016/0006-2952(88)90108-6. [DOI] [PubMed] [Google Scholar]
  26. Raunio H., Syngelmä T., Pasanen M., Juvonen R., Honkakoski P., Kairaluoma M. A., Sotaniemi E., Lang M. A., Pelkonen O. Immunochemical and catalytical studies on hepatic coumarin 7-hydroxylase in man, rat, and mouse. Biochem Pharmacol. 1988 Oct 15;37(20):3889–3895. doi: 10.1016/0006-2952(88)90070-6. [DOI] [PubMed] [Google Scholar]
  27. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  28. Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
  29. Santisteban I., Povey S., Shephard E. A., Phillips I. R. The major phenobarbital-inducible cytochrome P-450 gene subfamily (P450IIB) mapped to the long arm of human chromosome 19. Ann Hum Genet. 1988 May;52(Pt 2):129–135. doi: 10.1111/j.1469-1809.1988.tb01088.x. [DOI] [PubMed] [Google Scholar]
  30. Simmons D. L., Kasper C. B. Genetic polymorphisms for a phenobarbital-inducible cytochrome P-450 map to the Coh locus in mice. J Biol Chem. 1983 Aug 25;258(16):9585–9588. [PubMed] [Google Scholar]
  31. Skow L. C. Genetic variation for prolidase (PEP-4) in the mouse maps near the gene for glucosephosphate isomerase (GPI-1) on chromosome 7. Biochem Genet. 1981 Aug;19(7-8):695–700. doi: 10.1007/BF00484002. [DOI] [PubMed] [Google Scholar]
  32. Song B. J., Gelboin H. V., Park S. S., Yang C. S., Gonzalez F. J. Complementary DNA and protein sequences of ethanol-inducible rat and human cytochrome P-450s. Transcriptional and post-transcriptional regulation of the rat enzyme. J Biol Chem. 1986 Dec 15;261(35):16689–16697. [PubMed] [Google Scholar]
  33. Southern E. M., Anand R., Brown W. R., Fletcher D. S. A model for the separation of large DNA molecules by crossed field gel electrophoresis. Nucleic Acids Res. 1987 Aug 11;15(15):5925–5943. doi: 10.1093/nar/15.15.5925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  35. Spurr N. K., Gough A. C., Stevenson K., Wolf C. R. The human cytochrome P450 CYP3 locus: assignment to chromosome 7q22-qter. Hum Genet. 1989 Jan;81(2):171–174. doi: 10.1007/BF00293896. [DOI] [PubMed] [Google Scholar]
  36. Squires E. J., Negishi M. Reciprocal regulation of sex-dependent expression of testosterone 15 alpha-hydroxylase (P-450(15 alpha)) in liver and kidney of male mice by androgen. Evidence for a single gene. J Biol Chem. 1988 Mar 25;263(9):4166–4171. [PubMed] [Google Scholar]
  37. Staden R. The current status and portability of our sequence handling software. Nucleic Acids Res. 1986 Jan 10;14(1):217–231. doi: 10.1093/nar/14.1.217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Wainwright B. J., Watson E. K., Shephard E. A., Phillips I. R. RFLP for a human cytochrome P-450 gene at 19q13.1-qter (HGM8 provisional designation CYPI). Nucleic Acids Res. 1985 Jun 25;13(12):4610–4610. doi: 10.1093/nar/13.12.4610. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wolf C. R., Seilman S., Oesch F., Mayer R. T., Burke M. D. Multiple forms of cytochrome P-450 related to forms induced marginally by phenobarbital. Differences in structure and in the metabolism of alkoxyresorufins. Biochem J. 1986 Nov 15;240(1):27–33. doi: 10.1042/bj2400027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wood A. W., Taylor B. A. Genetic regulation of coumarin hydroxylase activity in mice. Evidence for single locus control on chromosome. J Biol Chem. 1979 Jul 10;254(13):5647–5651. [PubMed] [Google Scholar]

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