S-nitrosylation of Bcl-2 inhibits its ubiquitin-proteasomal degradation. A novel antiapoptotic mechanism that suppresses apoptosis.
Bcl-2 is a key apoptosis regulatory protein of the mitochondrial death pathway whose function is dependent on its expression levels. Although Bcl-2 expression is controlled by various mechanisms, post-translational modifications, such as ubiquitination and proteasomal degradation, have emerged as important regulators of Bcl-2 function. However, the underlying mechanisms of this regulation are unclear. We report here that Bcl-2 undergoes S-nitrosylation by endogenous nitric oxide (NO) in response to multiple apoptotic mediators and that this modification inhibits ubiquitin-proteasomal degradation of Bcl-2. Inhibition of NO production by the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide and by NO synthase inhibitor aminoguanidine effectively inhibited S-nitrosylation of Bcl-2, increased its ubiquitination, and promoted apoptotic cell death induced by chromium (VI). In contrast, the NO donors dipropylenetriamine NONOate and sodium nitroprusside showed opposite effects. The effect of NO on Bcl-2 stability was shown to be independent of its dephosphorylation. Mutational analysis of Bcl-2 further showed that the two cysteine residues of Bcl-2 (Cys158 and Cys229) are important in the S-nitrosylation process and that mutations of these cysteines completely inhibited Bcl-2 S-nitrosylation. Treatment of the cells with other stress inducers, including Fas ligand and buthionine sulfoxide, also induced Bcl-2 S-nitrosylation, suggesting that this is a general phenomenon that regulates Bcl-2 stability and function under various stress conditions. These findings indicate a novel function of NO and its regulation of Bcl-2, which provides a key mechanism for the control of apoptotic cell death and cancer development.
Azad
Neelam
N
Department of Pharmaceutical Sciences, West Virginia University, Morgantown, West Virginia 26506, USA.
Vallyathan
Val
V
Wang
Liying
L
Tantishaiyakul
Vimon
V
Stehlik
Christian
C
Leonard
Stephen S
SS
Rojanasakul
Yon
Y
D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D002840:Chromates / Q000494:pharmacology; D005434:Flow Cytometry; D006801:Humans; D047468:Immunoprecipitation; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009569:Nitric Oxide / Q000494:pharmacology*; D009603:Nitroso Compounds / Q000378:metabolism*; D010957:Plasmids; D061988:Proteasome Inhibitors; D011499:Protein Processing, Post-Translational; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D014452:Ubiquitins / Q000037:antagonists & inhibitors / Q000378:metabolism*
D002840:Chromates; D009603:Nitroso Compounds; D061988:Proteasome Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D014452:Ubiquitins; D009569:Nitric Oxide; C016104:sodium bichromate; D020169:Caspases