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Characterization of the CYP2C5 gene in 21L III/J rabbits. Allelic variation affects the expression of P450IIC5

Academic Article
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Overview

authors

  • Pendurthi, U. R.
  • Lamb, J. G.
  • Nguyen, N.
  • Johnson, Eric
  • Tukey, R. H.

publication date

  • August 1990

journal

  • Journal of Biological Chemistry  Journal

abstract

  • Rabbits exhibit two heritable phenotypes, 21L and 21H, with the latter displaying roughly 10-fold higher liver microsomal progesterone 21-hydroxylase activity. The higher activity of the 21H animals reflects the elevated expression of cytochrome P-450 1 (P450IIC5). Breeding studies indicate that this phenotypic difference is linked to allelic CYP2C5 genes (Johnson, E. F., Finlayson, M., Husjsak, C. M., Pendurthi, U. R., and Tukey, R. H. (1989) Arch. Biochem. Biophys. 273, 273-280). In this study, we report on the expression and structure of the CYP2C5 gene in the 21L inbred strain III/J. A cDNA encoding P450IIC5 was generated from III/J rabbit liver mRNA using polymerase chain reaction to amplify this sequence. DNA sequence analysis revealed that the cDNA encoded the same protein as the cDNA cloned previously from a 21H rabbit, demonstrating that structural differences in P450IIC5 do not underlie the phenotypic difference in the expression of 21-hydroxylase activity. DNA sequence analysis of the exons and surrounding intron regions in two III/J genomic clones that encode portions of the CYP2C5 gene showed that the exon sequences were identical to that of the P450IIC5 mRNA and that the codon positions of the exon/intron junctions are conserved with other class II genes. Southern blot analysis of DNA digested with BamHI and probed with a portion of DNA isolated from intron 5 revealed that the gene characterized in this study is an allele of the gene conferring the 21H phenotype. Primer extension analysis with a P450IIC5-specific oligonucleotide demonstrated that the CYP2C5 gene uses several transcriptional start sites in both 21H and 21L rabbits. Analysis of mRNA from livers of 21H and 21L rabbits with the P450IIC5-specific oligonucleotide showed a greater than 20-fold difference in the relative abundance of mRNA, which accounts quantitatively for the phenotypic difference in progesterone 21-hydroxylase activity.

subject areas

  • Alleles
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Blotting, Southern
  • Cloning, Molecular
  • Cytochrome P-450 Enzyme System
  • DNA
  • Female
  • Gene Expression Regulation, Enzymologic
  • Genes
  • Genetic Variation
  • Liver
  • Male
  • Molecular Sequence Data
  • Oligonucleotide Probes
  • Polymerase Chain Reaction
  • RNA, Messenger
  • Rabbits
  • Restriction Mapping
  • Sequence Homology, Nucleic Acid
  • Steroid 21-Hydroxylase
  • Steroid Hydroxylases
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Identity

International Standard Serial Number (ISSN)

  • 0021-9258

PubMed ID

  • 2387874
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Additional Document Info

start page

  • 14662

end page

  • 14668

volume

  • 265

issue

  • 24

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