Cytochrome P450CYP2J9, a new mouse arachidonic acid omega-1 hydroxylase predominantly expressed in brain

A cDNA encoding a new cytochrome P450 was isolated from a mouse brain libra ry. Sequence analysis reveals that this 1,958-base pair cDNA encodes a 57-5 8-kDa 502-amino acid polypeptide that is 70-91% identical to CYP2J subfamil y P450s and is designated CYP2J9. Recombinant CYP2J9 was co-expressed with NADPH-cytochrome P450 oxidoreductase (CYPOR) in Sf9 cells using a baculovir us system. Microsomes of CYP2J9/CYPOR-transfected cells metabolize arachido nic acid to 19-hydroxyeicosatetraenoic acid (HETE) thus CYP2J9 is enzymolog ically distinct from other P450s. Northern analysis reveals that CYP2J9 tra nscripts are present at high levels in mouse brain. Mouse brain microsomes biosynthesize 19-HETE. RNA polymerase chain reaction analysis demonstrates that CYP2J9 mRNAs are widely distributed in brain and most abundant in the cerebellum. Immunoblotting using an antibody raised against human CYP2J2 th at cross-reacts with CYP2J9 detects a 56-kDa protein band that is expressed in cerebellum and other brain segments and is regulated during postnatal d evelopment. In situ hybridization of mouse brain sections with a CYP2J9-spe cific riboprobe and immunohistochemical staining with the anti-human CYP2J2 IgG reveals abundant CYP2J9 mRNA and protein in cerebellar Purkinje cells. Importantly, 19-HETE inhibits the activity of recombinant P/Q-type Ca2+ ch annels that are known to be expressed preferentially in cerebellar Purkinje cells and are involved in triggering neurotransmitter release. Based on th ese data, we conclude that CYP2J9 is a developmentally regulated P450 that is abundant in brain, localized to cerebellar Purkinje cells, and active in the biosynthesis of 19-HETE, an eicosanoid that inhibits activity of P/Q-t ype Ca2+ channels. We postulate that CYP2J9 arachidonic acid products play important functional roles in the brain.