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Antioxid Redox Signal. 2015 Nov 10;23(14):1092-105. doi: 10.1089/ars.2014.6025. Epub 2015 Jan 9.

Targeting of Gamma-Glutamyl-Cysteine Ligase by miR-433 Reduces Glutathione Biosynthesis and Promotes TGF-β-Dependent Fibrogenesis.

Author information

1
1 Departamento de Biología Celular e Inmunología, Centro de Biología Molecular "Severo Ochoa, " Consejo Superior de Investigaciones Científicas-Universidad Autónoma de Madrid , Madrid, Spain .
2
2 Cellular Biology in Renal Diseases Laboratory, Universidad Autonoma de Madrid , Madrid, Spain .
3
3 Department of Metabolomics, CIC bioGUNE, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (Ciberehd) , Bizkaia, Spain .
4
4 Vascular Biology and Therapeutics Program, Department of Comparative Medicine, Yale University School of Medicine , New Haven, Connecticut.

Abstract

AIMS:

Glutathione (GSH) is the main antioxidant against cell damage. Several pathological states course with reduced nucleophilic tone and perturbation of redox homeostasis due to changes in the 2GSH/GSSG ratio. Here, we investigated the regulation of the rate-limiting GSH biosynthetic heterodimeric enzyme γ-glutamyl-cysteine ligase (GCL) by microRNAs (miRNAs).

RESULTS:

"In silico" analysis of the 3'- untranslated regions (UTRs) of both catalytic (GCLc) and regulatory (GCLm) subunits of GCL enabled an identification of miR-433 as a strong candidate for the targeting of GCL. Transitory overexpression of miR-433 in human umbilical vein endothelial cells (HUVEC) showed a downregulation of both GCLc and GCLm in a nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-independent manner. Increases in pro-oxidant stimuli such as exposure to hydrogen peroxide or GSH depletion in endothelial and hepatic cells caused an expected increase in GCLc and GCLm protein expression and abrogation of miR-433 levels, thus supporting a cross-regulation of these pathways. Treatment of HUVEC with miR-433 resulted in reduced antioxidant and redox potentials, increased S-glutathionylation, and reduced endothelial nitric oxide synthase activation. In vivo models of renal and hepatic fibrosis were associated with transforming growth factor β1 (TGF-β1)-related reduction of GCLc and GCLm levels that were miR-433 dependent.

INNOVATION AND CONCLUSION:

We describe for the first time an miRNA, miR-433, capable of directly targeting GCL and promoting functional consequences in endothelial physiology and fibrotic processes by decreasing GSH levels.

PMID:
25353619
PMCID:
PMC4657521
DOI:
10.1089/ars.2014.6025
[Indexed for MEDLINE]
Free PMC Article

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