Physiological role of the N-terminal processed P4501A1 targeted to mitochondria in erythromycin metabolism and reversal of erythromycin-mediated inhibition of mitochondrial protein synthesis.
Recently, we showed that the major species of beta-naphthoflavone-inducible rat liver mitochondrial P450MT2 consists of N-terminal truncated microsomal P4501A1 (+33/1A1) and that the truncated enzyme exhibits different substrate specificity as compared with intact P4501A1. The results of the present study show that P450MT2 targeted to COS cell mitochondria by transient transfection of P4501A1 cDNA is localized inside the mitochondrial inner membrane in a membrane-extrinsic orientation. Co-expression with wild type P4501A1 and adrenodoxin (Adx) cDNAs resulted in 5-7-fold higher erythromycin N-demethylation (ERND) in the mitochondrial fraction but minimal changes in the microsomal fraction of transfected cells. Erythromycin, a potent inhibitor of bacterial and mitochondrial protein synthesis, caused 8-12-fold higher accumulation of CYP1A1 mRNA, preferential accumulation of P450MT2, and 5-6-fold higher ERND activity in the mitochondrial compartment of rat C6 glioma cells. Consistent with the increased mitochondrial ERND activity, co-expression with P4501A1 and Adx in COS cells rendered complete protection against erythromycin-mediated mitochondrial translation inhibition. Mutations that specifically affect the mitochondrial targeting of P4501A1 also abolished protection against mitochondrial translation inhibition. These results for the first time suggest a physiological function for the xenobiotic inducible cytochrome P4501A1 against drug-mediated mitochondrial toxicity.
Anandatheerthavarada
H K
HK
Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6047, USA.
Vijayasarathy
C
C
Bhagwat
S V
SV
Biswas
G
G
Mullick
J
J
Avadhani
N G
NG
D000818:Animals; D019363:Cytochrome P-450 CYP1A1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004917:Erythromycin / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D008930:Mitochondria, Liver / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity; D019324:beta-Naphthoflavone / Q000494:pharmacology
D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D019324:beta-Naphthoflavone; D004917:Erythromycin; D019363:Cytochrome P-450 CYP1A1