Abstract
BackgroundSoluble guanylyl cyclase (sGC) plays a central role in nitric oxide (NO)-mediated signal transduction in the cardiovascular, nervous and gastrointestinal systems. Alternative RNA splicing has emerged as a potential mechanism to modulate sGC expression and activity. C-α1 sGC is an alternative splice form that is resistant to oxidation-induced protein degradation and demonstrates preferential subcellular distribution to the oxidized environment of endoplasmic reticulum (ER).Methodology/Principal FindingsHere we report that splicing of C-α1 sGC can be modulated by H2O2 treatment in BE2 neuroblastoma and MDA-MD-468 adenocarcinoma human cells. In addition, we show that the H2O2 treatment of MDA-MD-468 cells selectively decreases protein levels of PTBP1 and hnRNP A2/B1 splice factors identified as potential α1 gene splicing regulators by in silico analysis. We further demonstrate that down-regulation of PTBP1 by H2O2 occurs at the protein level with variable regulation observed in different breast cancer cells.Conclusions/SignificanceOur data demonstrate that H2O2 regulates RNA splicing to induce expression of the oxidation-resistant C-α1 sGC subunit. We also report that H2O2 treatment selectively alters the expression of key splicing regulators. This process might play an important role in regulation of cellular adaptation to conditions of oxidative stress.
Highlights
Alternative splicing expands transcriptome diversity [1,2] and allows cells to meet the requirements of an ever-changing extracellular environment
In this report we demonstrate that the oxidative stress induced by H2O2 influences splicing of the a1 Soluble guanylyl cyclase (sGC) gene (GUCY1A3) and selectively decreases protein level of PTBP1 and heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 splicing factors
We previously established that GUCY1A3 transcripts undergo alternative splicing and that the C-a1 sGC splice isoform encodes a protein that is resistant to ODQ-induced degradation [19,37]
Summary
Alternative splicing expands transcriptome diversity [1,2] and allows cells to meet the requirements of an ever-changing extracellular environment. Common stressors such as heat-shock, amino acid starvation or ethanol toxicity have been demonstrated to regulate alternative splicing [3,4]. Oxidative stress often persists in cellular microenvironment when there is an imbalance in the production and elimination of reactive oxygen species (ROS). This imbalance is associated with a plethora of pathologic conditions including carcinogenesis, cardiovascular disorders and neurodegeneration [5,6,7,8,9]. C-a1 sGC is an alternative splice form that is resistant to oxidation-induced protein degradation and demonstrates preferential subcellular distribution to the oxidized environment of endoplasmic reticulum (ER)
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