TY - JOUR
T1 - Redox Regulation of Human Rac1 Stability by the Proteasome in Human Aortic Endothelial Cells
AU - Kovacic, Hervé N.
AU - Irani, Kaikobad
AU - Goldschmidt-Clermont, Pascal J.
N1 - Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.
PY - 2001/12/7
Y1 - 2001/12/7
N2 - Rac1 has been shown to activate a NADPH oxidase complex producing superoxide anions in a variety of mammalian cell types. We evaluated the impact of Rac1-induced reactive oxygen species production on the turnover of Rac1 itself in human aortic endothelial cells. The concentration of a constitutively active mutant of Rac1 (Rac1V12) was increased by treatment of the cells with diphenylene iodinium (DPI), an inhibitor of the NADPH oxidase. Such an effect was not observed for the dominant negative form of Rac1 (Rac1 N17). We showed a decrease in proteolytic degradation of Rac1 V12 in the presence of DPI, and showed that short term treatment with H2O2 reverses the effect of DPI. We found that proteasome inhibitors (lactacystin and MG132) increased Rac1V12 protein level. In support of this finding, we have identified in the primary sequence of Rac1 a potential destruction box domain, which is known to be a signal for protein degradation mediated by the ubiquitin/proteasome system. We show that Rac1V12 is ubiquitinated before degradation. By contrast Rac1N17 induces an accumulation of the ubiquitinated form of Rac1. These results suggest that Rac1 activation of NADPH oxidase is necessary for the proteolytic degradation of Rac1 itself.
AB - Rac1 has been shown to activate a NADPH oxidase complex producing superoxide anions in a variety of mammalian cell types. We evaluated the impact of Rac1-induced reactive oxygen species production on the turnover of Rac1 itself in human aortic endothelial cells. The concentration of a constitutively active mutant of Rac1 (Rac1V12) was increased by treatment of the cells with diphenylene iodinium (DPI), an inhibitor of the NADPH oxidase. Such an effect was not observed for the dominant negative form of Rac1 (Rac1 N17). We showed a decrease in proteolytic degradation of Rac1 V12 in the presence of DPI, and showed that short term treatment with H2O2 reverses the effect of DPI. We found that proteasome inhibitors (lactacystin and MG132) increased Rac1V12 protein level. In support of this finding, we have identified in the primary sequence of Rac1 a potential destruction box domain, which is known to be a signal for protein degradation mediated by the ubiquitin/proteasome system. We show that Rac1V12 is ubiquitinated before degradation. By contrast Rac1N17 induces an accumulation of the ubiquitinated form of Rac1. These results suggest that Rac1 activation of NADPH oxidase is necessary for the proteolytic degradation of Rac1 itself.
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U2 - 10.1074/jbc.M107925200
DO - 10.1074/jbc.M107925200
M3 - Article
C2 - 11585836
AN - SCOPUS:0035824573
VL - 276
SP - 45856
EP - 45861
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 49
ER -