Related Experiment Videos
Nitric oxide down-regulates MKP-3 mRNA levels: involvement in endothelial cell protection from apoptosis
L Rössig1, J Haendeler, C Hermann
1Molecular Cardiology, Department of Internal Medicine IV, University of Frankfurt, Germany.
Abstract:
MAP kinase-dependent phosphorylation processes have been shown to interfere with the degradation of the antiapoptotic protein Bcl-2. The cytosolic MAP kinase phosphatase MAP kinase phosphatase-3 (MKP-3) induces apoptosis of endothelial cells in response to tumor necrosis factor alpha (TNFalpha) via dephosphorylation of the MAP kinase ERK1/2, leading to Bcl-2 proteolysis. Here we report that the endothelial cell survival factor nitric oxide (NO) down-regulated MKP-3 by destabilization of MKP-3 mRNA. This effect of NO was paralleled by a decrease in MKP-3 protein levels. Moreover, ERK1/2 was found to be protected against TNFalpha-induced dephosphorylation by coincubation of endothelial cells with the NO donor. Subsequently, both the decrease in Bcl-2 protein levels and the mitochondrial release of cytochrome c in response to TNFalpha were largely prevented by exogenous NO. In cells overexpressing MKP-3, no differences in phosphatase activity in the presence or absence of NO were found, excluding potential posttranslational modifications of MKP-3 protein by NO. These data demonstrate that upstream of the S-nitrosylation of caspase-3, NO exerts additional antiapoptotic effects in endothelial cells, which rely on the down-regulation of MKP-3 mRNA.
Insights
Nitric oxide (NO) protects endothelial cells from tumor necrosis factor alpha (TNFα)-induced apoptosis by reducing MAP kinase phosphatase-3 (MKP-3) mRNA stability. This NO-mediated mechanism prevents Bcl-2 degradation and cytochrome c release, highlighting a novel antiapoptotic pathway.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- MAP kinase (MAPK) signaling pathways regulate cell survival and apoptosis.
- MAP kinase phosphatase-3 (MKP-3) dephosphorylates and inactivates MAPK, promoting apoptosis by enabling Bcl-2 degradation.
- Tumor necrosis factor alpha (TNFα) induces endothelial cell apoptosis via MAPK activation.
Purpose of the Study:
- To investigate the role of nitric oxide (NO) in regulating MKP-3 expression and its impact on endothelial cell apoptosis.
- To elucidate the mechanism by which NO affects MKP-3 and its downstream targets.
Main Methods:
- Endothelial cells were treated with TNFα and NO donors.
- MKP-3 mRNA and protein levels were assessed.
- ERK1/2 phosphorylation, Bcl-2 degradation, and cytochrome c release were measured.
- Cells overexpressing MKP-3 were used to assess posttranslational modifications.
Main Results:
- NO down-regulated MKP-3 expression by destabilizing MKP-3 mRNA.
- NO protected ERK1/2 from TNFα-induced dephosphorylation.
- NO prevented TNFα-induced decrease in Bcl-2 protein levels and cytochrome c release.
- NO did not alter MKP-3 phosphatase activity, ruling out posttranslational modification.
Conclusions:
- NO exerts antiapoptotic effects in endothelial cells by down-regulating MKP-3 mRNA.
- This mechanism involves preventing Bcl-2 proteolysis and cytochrome c release, acting upstream of caspase-3 S-nitrosylation.
- NO represents a significant survival factor for endothelial cells through modulation of MAPK signaling.