Nitric oxide-dependent proteasomal degradation of cytochrome P450 2B proteins.
Exposure to inflammatory agents or cytokines causes the suppression of cytochrome P450 (CYP) enzyme activities and expression in liver and primary hepatocyte cultures. We showed previously that phenobarbital-induced CYP2B protein is down-regulated in primary cultures of rat hepatocytes after exposure to bacterial endotoxin (lipopolysaccharide) in a nitric oxide (NO) -dependent manner. In this study, we found that CYP2B proteins in primary rat hepatocyte cultures were suppressed >60% after 6 h of treatment with interleukin-1beta (IL-1). This effect was NO-dependent, and treatment of cells with the NO donors (Z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl) aminodiazen-1-ium-1,2-diolate (NOC-18), S-nitrosoglutathione, and S-nitroso-N-acetylpenicillamine also suppressed CYP2B proteins. However, the down-regulation by IL-1 was insensitive to inhibition of cGMP-dependent protein kinases. The down-regulation by IL-1 or NO donors was abolished by treatments with the proteasome inhibitors MG132 and lactacystin that did not affect NO production. The calpain inhibitor E64-d or the lysosomal protease inhibitors NH(4)Cl and chloroquine did not attenuate the down-regulation of CYP2B by IL-1. Treatment of HeLa cells expressing c-Myc-tagged CYP2B1 with NOC-18 down-regulated its expression and enhanced its ubiquitination. Treatment of rat liver microsomes with S-nitrosoglutathione caused S-nitrosylation of CYP2B protein and enhanced the ubiquitination pattern of CYP2B compared with unmodified CYP2B in an in vitro ubiquitination assay. These data are consistent with the hypothesis that NO-dependent CYP2B ubiquitination and proteasomal degradation are dependent on protein modification by reactive nitrogen species.
Lee
Choon-Myung
CM
Department of Pharmacology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Kim
Bong-Yoon
BY
Li
Lian
L
Morgan
Edward T
ET
D000818:Animals; D019362:Cytochrome P-450 CYP2B1 / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000201:enzymology*; D007375:Interleukin-1 / Q000494:pharmacology; D015850:Interleukin-6 / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009569:Nitric Oxide / Q000494:pharmacology*; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014158:Transcription, Genetic
D007375:Interleukin-1; D015850:Interleukin-6; D009569:Nitric Oxide; D003577:Cytochrome P-450 Enzyme System; D019362:Cytochrome P-450 CYP2B1; D046988:Proteasome Endopeptidase Complex