Mitochondria-targeted cytochrome P450 2E1 induces oxidative damage and augments alcohol-mediated oxidative stress.
The ethanol-inducible cytochrome P450 2E1 (CYP2E1) is also induced under different pathological and physiological conditions. Studies including ours have shown that CYP2E1 is bimodally targeted to both the endoplasmic reticulum (microsomes) (mc CYP2E1) and mitochondria (mt CYP2E1). In this study we investigated the role of mtCYP2E1 in ethanol-mediated oxidative stress in stable cell lines expressing predominantly mt CYP2E1 or mc CYP2E1. The ER+ mutation (A2L, A9L), which increases the affinity of the nascent protein for binding to the signal recognition particle, preferentially targets CYP2E1 to the endoplasmic reticulum. The Mt+ (L17G) and Mt++ (I8R, L11R, L17R) mutant proteins, showing progressively lower affinity for signal recognition particle binding, were targeted to mitochondria at correspondingly higher levels. The rate of GSH depletion, used as a measure of oxidative stress, was higher in cells expressing Mt++ and Mt+ proteins as compared with cells expressing ER+ protein. In addition, the cellular level of F(2)-isoprostanes, a direct indicator of oxidative stress, was increased markedly in Mt++ cells after ethanol treatment. Notably, expression of Mt++ CYP2E1 protein in yeast cells caused more severe mitochondrial DNA damage and respiratory deficiency than the wild type or ER+ proteins as tested by the inability of cells to grow on glycerol or ethanol. Additionally, liver mitochondria from ethanol-fed rats containing high mt CYP2E1 showed higher levels of F(2)-isoprostane production. These results strongly suggest that mt CYP2E1 induces oxidative stress and augments alcohol-mediated cell/tissue injury.
Department of Animal Biology and the Mari Lowe Center for Comparative Oncology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Seema
S
Bansal
Chuan-Peng
CP
Liu
Naresh B V
NB
Sepuri
Hindupur K
HK
Anandatheerthavarada
Venkatesh
V
Selvaraj
Jan
J
Hoek
Ginger L
GL
Milne
F Peter
FP
Guengerich
Narayan G
NG
Avadhani
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D019392:Cytochrome P-450 CYP2E1 / Q000378:metabolism*; D004272:DNA, Mitochondrial / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D000431:Ethanol / Q000737:chemistry; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism; D018384:Oxidative Stress; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000378:metabolism
D004272:DNA, Mitochondrial; D000431:Ethanol; D019392:Cytochrome P-450 CYP2E1; D005978:Glutathione