Sequence of a novel cytochrome CYP2B cDNA coding for a protein which is expressed in a sebaceous gland, but not in the liver

Abstract

The major phenobarbital-inducible rat hepatic cytochromes P-450, CYP2B1 and CYP2B2, are the paradigmatic members of a cytochrome P-450 gene subfamily that contains at least seven additional members. Specific oligonucleotide probes for these genomic members of the CYP2B subfamily were used to assess their tissue-specific expression. In Northern-blot analysis a probe specific to gene 4 (which is designated now as CYP2B12) hybridized to a single mRNA present in the preputial gland, an organ which is used as a model for sebaceous glands, but did not hybridize to mRNA isolated from the liver or from five other tissues of untreated or Aroclor 1254-treated rats. The cDNA sequence for the CYP2B12 RNA was determined from overlapping cDNA clones and contained a long open reading frame of 1476 bp. The nucleotide sequence of the CYP2B12 cDNA was 85% similar to the sequence of the CYP2B1 cDNA in its coding region and was different from any CYP2B cDNA characterized until now. The cDNA-derived primary structure of the CYP2B12 protein contains a signal sequence for its insertion into the endoplasmic reticulum and the putative haem-binding site characteristic of cytochromes P-450. A part of the potential haem pocket of CYP2B12 was identical with a similar structure in a bacterial protocatechuate dioxygenase. In immunoblot analysis of preputial-gland microsomes, antibodies against CYP2B1 recognized a single abundant protein with a lower apparent molecular mass than that of CYP2B1. Our results demonstrate that the CYP2B12 protein has the potential to be enzymically active and are the first demonstration that a member of the CYP2B subfamily is expressed exclusively and at high levels in an extrahepatic organ.

Differentiating Keratinocytes Express a Novel Cytochrome P450 Enzyme, CYP2B19, Having Arachidonate Monooxygenase Activity

“…Degener- ate oligonucleotides specific for mammalian CYP2B genes amplified a fragment of a novel gene, Cyp2b19, with RNA isolated from developing limbs (E15.5). This nucleotide sequence was more closely related to two rat genes, CYP2B12 (7,8) and CYP2B15 (24), than to previously identified murine genes. Authenticity of this partial cDNA was confirmed by the specific hybridization of antisense CYP2B19 cRNA to fetal mouse epidermis (E15.5; Fig.…”

“…CYP2B12, a homologous cytochrome P450 in rat skin, is also an arachidonic acid monooxygenase. Like CYP2B19 it is keratinocyte-specific but generates only EETs from arachidonate (7,8). Expression data indicate that the catalytic activities of these two enzymes are highly compartmentalized to the most differentiated, viable cell layers in the epidermis (granular cells), hair follicles (inner root sheath), or sebaceous glands (sebocytes).…”

A Keratinocyte-specific Epoxygenase, CYP2B12, Metabolizes Arachidonic Acid with Unusual Selectivity, Producing a Single Major Epoxyeicosatrienoic Acid

“…Originally called gene 4, it was one of several partial genomic clones that proved the multiplicity of the CYP2B gene subfamily. Gene 4 expression was not detected in the liver or other organs but rather in preputial glands (4), the large, paired sebaceous glands beneath the genital skin in rodents. This finding led Friedberg et al (4) to isolate and characterize a full-length cDNA corresponding to gene 4 from a preputial gland cDNA library.…”

Molecular Biological Methods for Characterizing Drug-Metabolizing Enzymes in Hepatic and Extrahepatic Tissues

“…However, it was not known whether these clones represent pseudogens and in which tissues the functional genes would be expressed. We have isolated RNAs from several tissues and have hybridized them to oligonucleotide probes which were specific to only one of the genomic clones isolated by Adesnik and co-workers [35]. As expected, the CYP2B1-specific probe hybridized mainly to hepatic RNA isolated from phenobarbital- treated rats ( fig.…”