Abstract
The regulation of the synthesis of the endothelial-derived vasoconstrictor endothelin-1 (ET-1) is a complex process encompassing transcriptional as well as mRNA stability mechanisms. We have described recently the existence of a mechanism for the control of ET-1 expression based on the mRNA-destabilizing capacity of specific cytosolic proteins through interaction with AU-rich elements (AREs) present in the 3' untranslated region of the gene. We now identify glyceraldehyde-3'-phosphate dehydrogenase (GAPDH) as a protein which binds to the AREs and is responsible for the destabilization of the mRNA. Oxidant stress alters the binding of GAPDH to the mRNA and its capacity to modulate ET-1 expression, a phenomenon occurring through specific S glutathionylation of the catalytically active residue Cys 152. Finally, we provide data consistent with a role for GAPDH in mRNA unwinding, yielding this molecule more prone to degradation. In contrast, S-thiolated GAPDH appears unable to modify mRNA unwinding, thus facilitating enhanced stability. Taken together, these results describe a novel, redox-based mechanism regulating mRNA stability and add a new facet to the panoply of GAPDH cellular homeostatic actions.
Highlights
The endothelium contributes greatly to vascular homeostasis by functioning as a sensor of physical and chemical stimuli and responding to them with specific signals
We show that the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) constitutes the major component interacting with functional AU-rich elements (AREs) of the 3Ј untranslated region (3Ј UTR) of ET-1 mRNA and that this interaction is associated with reduced ET-1 expression by mRNA destabilization
We showed that endothelial ET-1 expression is under the control of a mechanism based on the mRNAdestabilizing capacity of specific cytosolic proteins through interaction with AREs present in the 3Ј UTR of the ET-1 gene
Summary
The endothelium contributes greatly to vascular homeostasis by functioning as a sensor of physical and chemical stimuli and responding to them with specific signals. We report the identification of RNA binding proteins interacting with AREs of ET-1 mRNA from vascular endothelial cells and the mechanism by which the regulation of the expression of the gene is achieved. This phenomenon occurs through specific posttranslational modification of the catalytically active GAPDH residue Cys 152 by the incorporation of glutathione, or S-glutathionylation We suggest that this mechanism plays a critical role in the regulation of ET-1 mRNA expression by oxidative or nitrosative agents. Such a mechanism of control may be of importance in the vascular environment, where oxidative stress is able to promote cardiovascular disease by alteration of the synthesis of vasoactive factors
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