Alternative Splicing of CYP2D mRNA in Human Breast Tissue

Abstract

The human cytochrome P450 (CYP) 2D subfamily comprises theCYP2D6gene and four pseudogenes,CYP2D7P1and2andCYP2D8P1and2.TheCYP2D6gene product is a prominent drug-metabolizing enzyme, which is probably constitutive and has no known inducing agents. Alternative splicing of the pre-mRNAs of these genes has been detected in human liver and breast tissue. RNA–PCR, competitive RNA–PCR, Southern blotting, cDNA sequencing, and gene-specific PCR have been used to fully characterize the alternatively spliced forms of CYP2D mRNA in human breast tissue in the region of exon 5 to 8. Such alternative splicing could regulate the expression of CYP2D6 protein. A full-length mRNA (exons 5 to 8), and variants c (exon 6 deleted), b′ (3′ portion of exon 6 deleted), e (3′ portion of exon 6 deleted, 3′ 57-bp portion of intron 6 included), d (3′ 57-bp portion of intron 6 included), and b (intron 6 included) were characterized and quantitated. Variant c was derived fromCYP2D6,variants d, e, and b were fromCYP2D7P,and variant b′ and full-length mRNA were derived from bothCYP2D6and2D7P.Full-length mRNA was a minor form in human breast tissue where variants b′ and c predominated. Human breast tumor MCF-7 cells had CYP2D mRNA splice variant patterns similar to those of human breast tissue, while human liver tumor HepG2 cells had wild-type mRNA predominating. These results suggest that CYP2D6 could be regulated tissue specifically using tissue-specific alternative mRNA splicing.