Publications (2)23.34 Total impact
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Article: Proteasome activation by hepatitis C core protein is reversed by ethanol-induced oxidative stress.
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ABSTRACT: The proteasome is a major cellular proteinase. Its activity is modulated by cellular oxidants. Hepatitis C core protein and ethanol exposure both cause enhanced oxidant generation. The aim was to investigate whether core protein, by its ability to generate oxidants, alters proteasome activity and whether these alterations are further affected by ethanol exposure. These interactions were examined in Huh-7 cell lines that expressed inducible HCV core protein and/or constitutive cytochrome P450 2E1 (CYP2E1) and as purified components in a cell-free system. Chymotrypsin-like proteasome activity was measured fluorometrically. Proteasome activity in core-positive 191-20 cells was 20% higher than that in core-negative cells and was enhanced 3-fold in CYP2E1-expressing L14 cells. Exposure of core-positive cells to glutathione ethyl ester, catalase, or the CYP2E1 inhibitor diallyl sulfide partially reversed the elevation of proteasome activity in core-positive cells, whereas ethanol exposure suppressed proteasome activity. The results indicate that proteasome activity was up-regulated by low levels of core-induced oxidative stress but down-regulated by high levels of ethanol-elicited stress. These findings were partially mimicked in a cell-free system. Addition of core protein enhanced the peptidase activity of purified 20S proteasome containing the proteasome activator PA28 and was further potentiated by addition of liver mitochondrial and/or microsome fractions. However, proteasome activation was significantly attenuated when fractions were obtained from ethanol-fed animals. HCV core protein interacts with PA28, mitochondrial, and endoplasmic reticulum proteins to cause low levels of oxidant stress and proteasome activation, which is dampened during ethanol metabolism when oxidant generation is higher.Gastroenterology 06/2008; 134(7):2144-52. · 11.68 Impact Factor -
Article: Peroxynitrite alters the catalytic activity of rodent liver proteasome in vitro and in vivo.
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ABSTRACT: The proteasome is an important multicatalytic enzyme complex that degrades misfolded and oxidized proteins, signal transduction factors, and antigenic peptides for presentation. We investigated the in vitro effects of peroxynitrite (PN) on the peptidase activity of both crude 20S and 26S and purified 20S proteasome preparations from rat liver as well as proteasome activity in Hep G2 cells and in mouse liver. Crude and purified proteasome preparations were exposed to PN or to the PN donor, 3-morpholinosydnonimine hydrochloride (SIN-1), and then assayed for chymotrypsin-like activity. For in vivo experiments, mice were treated with molsidomine, which is metabolized to SIN-1 in liver. PN and SIN-1 dose-dependently modulated the chymotrypsin-like activity of the 20S proteasome: lower concentrations enhanced proteasome activity, and higher concentrations caused its decline. The NO donor S-nitroso-N-acetylpenicillamine (SNAP), at all concentrations, suppressed 20S proteasome activity. We observed similar results when liver soluble fractions (S-100) were treated with PN, SIN-1, or SNAP, except that enzyme activity in S-100 fractions was less sensitive than the purified enzymes to these agents. Treatment of Hep G2 cells with 0.01 or 0.1 mmol/L SIN-1 stimulated in situ proteasome activity in these cells, while 1 mmol/L SIN-1 suppressed it. SNAP treatment did not affect proteasome activity in Hep G2 cells. Mice treated with molsidomine had enhanced liver proteasome activity 6 hours after treatment, but after 24 hours enzyme activity declined below control levels. In conclusion, PN dose-dependently modulated proteasome activity, regulating protein degradation by the proteasome in liver cells.Hepatology 10/2004; 40(3):574-82. · 11.66 Impact Factor
