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(Received for publication, July 10, 1996, and in revised form, December 30, 1996)
From the From the Laboratories of A cDNA encoding a P450 monooxygenase was
amplified from reverse transcribed rat heart and liver total RNA by
polymerase chain reaction using primers based on the 5
Volume 272, Number 19,
Issue of May 9, 1997
pp. 12551-12559
©1997 by The American Society for Biochemistry and Molecular Biology, Inc.
,
,
and
Pulmonary
Pathobiology, § Molecular Biophysics, and
¶ Experimental Pathology, NIEHS, National Institutes of Health,
Research Triangle Park, North Carolina 27709, the 
Department of
Pathology, Duke University Medical Center, Durham, North Carolina
27719, and the ** Department of Molecular Genetics, University of Texas
Southwestern Medical Center, Dallas, Texas 75235
- and 3-end
sequences of two rat pseudogenes, CYP2J3P1 and
CYP2J3P2. Sequence analysis revealed that this 1,778-base
pair cDNA contained an open reading frame and encoded a new 502 amino acid protein designated CYP2J3. Based on the deduced amino acid
sequence, CYP2J3 was approximately 70% homologous to both human CYP2J2
and rabbit CYP2J1. Recombinant CYP2J3 protein was co-expressed with
NADPH-cytochrome P450 oxidoreductase in Sf9 insect cells using a
baculovirus expression system. Microsomal fractions of
CYP2J3/NADPH-cytochrome P450 oxidoreductase-transfected cells
metabolized arachidonic acid to 14,15-, 11,12-, and
8,9-epoxyeicosatrienoic acids and 19-hydroxyeicosatetraenoic acid as
the principal reaction products (catalytic turnover, 0.2 nmol of
product/nmol of cytochrome P450/min at 37 °C). Immunoblotting of
microsomal fractions prepared from rat tissues using a polyclonal
antibody raised against recombinant CYP2J2 that cross-reacted with
CYP2J3 but not with other known rat P450s demonstrated abundant
expression of CYP2J3 protein in heart and liver. Immunohistochemical
staining of formalin-fixed paraffin-embedded rat heart tissue sections
using the anti-CYP2J2 IgG and avidin-biotin-peroxidase detection
localized expression of CYP2J3 primarily to atrial and ventricular
myocytes. In an isolated-perfused rat heart model, 20 min of global
ischemia followed by 40 min of reflow resulted in recovery of only
44 ± 6% of base-line contractile function. The addition of 5 µM 11,12-epoxyeicosatrienoic acid to the perfusate prior
to global ischemia resulted in a significant 1.6-fold improvement in
recovery of cardiac contractility (69 ± 5% of base line,
p = 0.01 versus vehicle alone).
Importantly, neither 14,15-epoxyeicosatrienoic acid nor
19-hydroxyeicosatetraenoic acid significantly improved functional
recovery following global ischemia, demonstrating the specificity of
the biological effect for the 11,12-epoxyeicosatrienoic acid
regioisomer. Based on these data, we conclude that (a)
CYP2J3 is one of the predominant enzymes responsible for the oxidation
of endogenous arachidonic acid pools in rat heart myocytes and
(b) 11,12-epoxyeicosatrienoic acid may play an important
functional role in the response of the heart to ischemia.
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P. Su, K. M. Kaushal, and D. L. Kroetz Inhibition of renal arachidonic acid omega -hydroxylase activity with ABT reduces blood pressure in the SHR Am J Physiol Regulatory Integrative Comp Physiol, August 1, 1998; 275(2): R426 - R438. [Abstract] [Full Text] [PDF]
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F. Cabello-Hurtado, Y. Batard, J.-P. Salaun, F. Durst, F. Pinot, and D. Werck-Reichhart Cloning, Expression in Yeast, and Functional Characterization of CYP81B1, a Plant Cytochrome P450 That Catalyzes In-chain Hydroxylation of Fatty Acids J. Biol. Chem., March 27, 1998; 273(13): 7260 - 7267. [Abstract] [Full Text] [PDF]
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D. C. Zeldin, J. Foley, S. M. Goldsworthy, M. E. Cook, J. E. Boyle, J. Ma, C. R. Moomaw, K. B. Tomer, C. Steenbergen, and S. Wu CYP2J Subfamily Cytochrome P450s in the Gastrointestinal Tract: Expression, Localization, and Potential Functional Significance Mol. Pharmacol., June 1, 1997; 51(6): 931 - 943. [Abstract] [Full Text]
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K. Node, X.-L. Ruan, J. Dai, S.-X. Yang, L. Graham, D. C. Zeldin, and J. K. Liao Activation of Galpha s Mediates Induction of Tissue-type Plasminogen Activator Gene Transcription by Epoxyeicosatrienoic Acids J. Biol. Chem., May 4, 2001; 276(19): 15983 - 15989. [Abstract] [Full Text] [PDF]
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W. Qu, J. A. Bradbury, C.-C. Tsao, R. Maronpot, G. J. Harry, C. E. Parker, L. S. Davis, M. D. Breyer, M. P. Waalkes, J. R. Falck, et al. Cytochrome P450 CYP2J9, a New Mouse Arachidonic Acid omega -1 Hydroxylase Predominantly Expressed in Brain J. Biol. Chem., June 29, 2001; 276(27): 25467 - 25479. [Abstract] [Full Text] [PDF]
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D. C. Zeldin Epoxygenase Pathways of Arachidonic Acid Metabolism J. Biol. Chem., September 21, 2001; 276(39): 36059 - 36062. [Full Text] [PDF]
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