Mouse Cyp4a isoforms: enzymatic properties, gender- and strain-specific expression, and role in renal 20-hydroxyeicosatetraenoic acid formation.
AA (arachidonic acid) hydroxylation to 20-HETE (20-hydroxyeicosatetraenoic acid) influences renal vascular and tubular function. To identify the CYP (cytochrome P450) isoforms catalysing this reaction in the mouse kidney, we analysed the substrate specificity of Cyp4a10, 4a12a, 4a12b and 4a14 and determined sex- and strain-specific expressions. All recombinant enzymes showed high lauric acid hydroxylase activities. Cyp4a12a and Cyp4a12b efficiently hydroxylated AA to 20-HETE with V(max) values of approx. 10 nmol x nmol(-1) x min(-1) and K(m) values of 20-40 microM. 20-Carboxyeicosatetraenoic acid occurred as a secondary metabolite. AA hydroxylase activities were approx. 25-75-fold lower with Cyp4a10 and not detectable with Cyp4a14. Cyp4a12a and Cyp4a12b also efficiently converted EPA (eicosapentaenoic acid) into 19/20-OH- and 17,18-epoxy-EPA. In male mice, renal microsomal AA hydroxylase activities ranged between approx. 100 (NMRI), 45-55 (FVB/N, 129 Sv/J and Balb/c) and 25 pmol x min(-1) x mg(-1) (C57BL/6). The activities correlated with differences in Cyp4a12a protein and mRNA levels. Treatment with 5alpha-dihydrotestosterone induced both 20-HETE production and Cyp4a12a expression more than 4-fold in male C57BL/6 mice. All female mice showed low AA hydroxylase activities (15-25 pmol x min(-1) x mg(-1)) and very low Cyp4a12a mRNA and protein levels, but high Cyp4a10 and Cyp4a14 expression. Renal Cyp4a12b mRNA expression was almost undetectable in both sexes of all strains. Thus Cyp4a12a is the predominant 20-HETE synthase in the mouse kidney. Cyp4a12a expression determines the sex- and strain-specific differences in 20-HETE generation and may explain sex and strain differences in the susceptibility to hypertension and target organ damage.
Muller
Dominik N
DN
Max Delbrueck Center for Molecular Medicine, Robert-Roessle-Str. 10, 13092 Berlin, Germany.
Schmidt
Cosima
C
Barbosa-Sicard
Eduardo
E
Wellner
Maren
M
Gross
Volkmar
V
Hercule
Hantz
H
Markovic
Marija
M
Honeck
Horst
H
Luft
Friedrich C
FC
Schunck
Wolf-Hagen
WH
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D000072470:Cytochrome P450 Family 4; D017931:DNA Primers; D005230:Fatty Acids, Nonesterified / Q000378:metabolism; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis*; D007527:Isoenzymes / Q000235:genetics; D007668:Kidney / Q000201:enzymology*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D013045:Species Specificity
D017931:DNA Primers; D005230:Fatty Acids, Nonesterified; D006893:Hydroxyeicosatetraenoic Acids; D007527:Isoenzymes; D012313:RNA; D003577:Cytochrome P-450 Enzyme System; C493210:Cyp4a12a protein, mouse; D000072470:Cytochrome P450 Family 4