Expression and alternative splicing of the cytochrome p-450 CYP2A7

S. Ding, B. G. Lake, T. Friedberg, C. R. Wolf

    Research output: Contribution to journalArticle

    87 Citations (Scopus)

    Abstract

    In order to investigate the relative levels of expression of human cytochrome P-450 (P-450) CYP2A genes and determine how this relates to polymorphism in coumarin hydroxylase activity, cDNA clones for members of the CYP2A gene family were isolated. These clones were CYP2A6, CYP2A7 and an alternatively spliced version of CYP2A7 (CYP2A7AS). The latter clone was missing exon 2, but contained a 10 bp segment of intron 1. Translation of CYP2A7AS resulted in an in-frame deletion of 51 amino acids. The expression of these cDNAs in COS-7 cells showed that both CYP2A6 and CYP2A7 generated a protein of molecular mass 49 kDa, whereas the protein product of CYP2A7AS was about 44 kDa. Only the CYP2A6 had coumarin hydroxylase activity. The relative level of CYP2A7 and CYP2A7AS mRNA was investigated by reverse transcription followed by PCR (RT-PCR) using human liver RNAs and an RNA sample from a human skin fibroblast cell line. In one of five liver RNAs studied, the aberrantly spliced CYP2A7 mRNA was 3-4-fold more abundant than the normal mRNA. The other samples contained very low levels of this mRNA species. Interestingly, CYP2A7AS mRNA was the major CYP2A7 mRNA detected in the fibroblast cell line. In this case only a protein band of 44 kDa was observed by Western-blot analysis. The relative level of mRNA encoding CYP2A6 and CYP2A7 was established in seven human liver samples by RT-PCR and found to range between 1:0.5 and 1:3. These data strengthen the previous findings that alternative splicing is an important factor in determining the levels of many human P-450s and that this may be subject to tissue-specific effects. Whether in this case the protein product has some function remains to be determined.

    Original languageEnglish
    Pages (from-to)161-166
    Number of pages6
    JournalBiochemical Journal
    Volume306
    Issue number1
    Publication statusPublished - 1995

    Fingerprint

    Alternative Splicing
    Cytochromes
    Messenger RNA
    Liver
    Clone Cells
    RNA
    Fibroblasts
    Polymerase Chain Reaction
    Proteins
    Complementary DNA
    Genes
    Cells
    Cell Line
    COS Cells
    Molecular mass
    Transcription
    human CYP2A7 protein
    Polymorphism
    Introns
    Cytochrome P-450 Enzyme System

    Cite this

    Ding, S. ; Lake, B. G. ; Friedberg, T. ; Wolf, C. R. / Expression and alternative splicing of the cytochrome p-450 CYP2A7. In: Biochemical Journal. 1995 ; Vol. 306, No. 1. pp. 161-166.
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    abstract = "In order to investigate the relative levels of expression of human cytochrome P-450 (P-450) CYP2A genes and determine how this relates to polymorphism in coumarin hydroxylase activity, cDNA clones for members of the CYP2A gene family were isolated. These clones were CYP2A6, CYP2A7 and an alternatively spliced version of CYP2A7 (CYP2A7AS). The latter clone was missing exon 2, but contained a 10 bp segment of intron 1. Translation of CYP2A7AS resulted in an in-frame deletion of 51 amino acids. The expression of these cDNAs in COS-7 cells showed that both CYP2A6 and CYP2A7 generated a protein of molecular mass 49 kDa, whereas the protein product of CYP2A7AS was about 44 kDa. Only the CYP2A6 had coumarin hydroxylase activity. The relative level of CYP2A7 and CYP2A7AS mRNA was investigated by reverse transcription followed by PCR (RT-PCR) using human liver RNAs and an RNA sample from a human skin fibroblast cell line. In one of five liver RNAs studied, the aberrantly spliced CYP2A7 mRNA was 3-4-fold more abundant than the normal mRNA. The other samples contained very low levels of this mRNA species. Interestingly, CYP2A7AS mRNA was the major CYP2A7 mRNA detected in the fibroblast cell line. In this case only a protein band of 44 kDa was observed by Western-blot analysis. The relative level of mRNA encoding CYP2A6 and CYP2A7 was established in seven human liver samples by RT-PCR and found to range between 1:0.5 and 1:3. These data strengthen the previous findings that alternative splicing is an important factor in determining the levels of many human P-450s and that this may be subject to tissue-specific effects. Whether in this case the protein product has some function remains to be determined.",
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    Ding, S, Lake, BG, Friedberg, T & Wolf, CR 1995, 'Expression and alternative splicing of the cytochrome p-450 CYP2A7', Biochemical Journal, vol. 306, no. 1, pp. 161-166.

    Expression and alternative splicing of the cytochrome p-450 CYP2A7. / Ding, S.; Lake, B. G.; Friedberg, T.; Wolf, C. R.

    In: Biochemical Journal, Vol. 306, No. 1, 1995, p. 161-166.

    Research output: Contribution to journalArticle

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    AU - Lake, B. G.

    AU - Friedberg, T.

    AU - Wolf, C. R.

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    N2 - In order to investigate the relative levels of expression of human cytochrome P-450 (P-450) CYP2A genes and determine how this relates to polymorphism in coumarin hydroxylase activity, cDNA clones for members of the CYP2A gene family were isolated. These clones were CYP2A6, CYP2A7 and an alternatively spliced version of CYP2A7 (CYP2A7AS). The latter clone was missing exon 2, but contained a 10 bp segment of intron 1. Translation of CYP2A7AS resulted in an in-frame deletion of 51 amino acids. The expression of these cDNAs in COS-7 cells showed that both CYP2A6 and CYP2A7 generated a protein of molecular mass 49 kDa, whereas the protein product of CYP2A7AS was about 44 kDa. Only the CYP2A6 had coumarin hydroxylase activity. The relative level of CYP2A7 and CYP2A7AS mRNA was investigated by reverse transcription followed by PCR (RT-PCR) using human liver RNAs and an RNA sample from a human skin fibroblast cell line. In one of five liver RNAs studied, the aberrantly spliced CYP2A7 mRNA was 3-4-fold more abundant than the normal mRNA. The other samples contained very low levels of this mRNA species. Interestingly, CYP2A7AS mRNA was the major CYP2A7 mRNA detected in the fibroblast cell line. In this case only a protein band of 44 kDa was observed by Western-blot analysis. The relative level of mRNA encoding CYP2A6 and CYP2A7 was established in seven human liver samples by RT-PCR and found to range between 1:0.5 and 1:3. These data strengthen the previous findings that alternative splicing is an important factor in determining the levels of many human P-450s and that this may be subject to tissue-specific effects. Whether in this case the protein product has some function remains to be determined.

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