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

Abstract

The CYP monooxygenase, CYP2B12, is the first identified skin-specific cytochrome P450 enzyme. It is characterized by high, constitutive expression in an extrahepatic tissue, the sebaceous glands of cutaneous tissues. It is expressed exclusively in a subset of differentiated keratinocytes called sebocytes, as demonstrated by Northern blot analysis, in situ hybridization, and polymerase chain reaction. The onset of its expression coincides with the morphological appearance of sebaceous glands in the neonatal rat. Recombinant CYP2B12 produced in Escherichia coli epoxidizes arachidonic acid to 11,12-and 8,9-epoxyeicosatrienoic acids (80 and 20% of total metabolites, respectively). The identification of arachidonic acid as a substrate for this skin-specific CYP monooxygenase suggests an endogenous function in keratinocytes in the generation of bioactive lipids and intracellular signaling.Mammalian CYP monooxygenases (CYP gene superfamily) oxidatively metabolize small, hydrophobic compounds, including steroids, sterols, fatty acids, fat-soluble vitamins, drugs, and toxins (1). These enzymes are expressed in most mammalian tissues, where they function in the biosynthesis or catabolism of endogenous and exogenous substrates. Members of the CYP2B gene subfamily are often called phenobarbital-inducible cytochrome P450 enzymes because phenobarbital treatment results in transcriptional activation of certain hepatic CYP2B genes (2). A typical CYP2B monooxygenase has 16-hydroxylase activity with androgenic steroids, such as testosterone, and is expressed in the liver and organs such as kidney, lung, and testis (2).CYP2B12 has long been known to be an unusual member of the CYP2B gene subfamily. The gene encoding this enzyme (CYP2B12) was discovered by Atchison and Adesnik (3). 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. More than 10 years after its discovery, the novel cytochrome P450 named CYP2B12 is now recognized as the first identified skin-specific cytochrome P450 enzyme.CYP2B12 has remained enigmatic not only for its restricted tissue-specific expression but also because its substrate was not identified. No activity could be demonstrated with preputial gland microsomes using typical CYP2B substrates, such as testosterone, androstenedione, benzphetamine, and ethoxy-, pentoxy-, and benzoyresorufin (2, 4). Our preliminary investigations of a novel CYP2B in mouse skin 1 having sequence homology to rat CYP2B12 renewed interest in this cutaneous P450 enzyme. We report here that CYP2B12 is a gene product unique to sebocytes, a subset of differentiated keratinocytes, and that arachidonate is a substrate for CYP2B12. It is hypothesiz...

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

“…All three genes were found to be uniquely expressed in differentiated, skin keratinocytes (Table II). Data for rat CYP2B12 have been reported (8). Because the expression patterns of CYP2B15 are identical to those of CYP2B19, the results in Fig.…”

“…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).…”

Human tear film and meibum. Very long chain wax esters and (O-acyl)-omega-hydroxy fatty acids of meibum

“…However, their enzymatic epoxidation through cytochrome P450, for example, is a possibility. CYP2B12 transcripts were found in the mouse skin and meibomian glands ( 32,33 ). Suzuki et al ( 33 ) studied the cytochrome P450 gene regulation by 17 ␤ -estradiol in mouse meibomian glands.…”

Eicosanoids in skin inflammation

“…Although some AA-specific mono-oxygenases have been identified in epidermal keratinocytes [86,87], the overall contribution of CYP in cutaneous eicosanoid production remains to be explored. …”