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Engineering Artificial Factors to Specifically Manipulate Alternative Splicing in Human Cells
Published on: April 26, 2017
Multiple effects of digoxin on subsets of cancer-associated genes through the alternative splicing pathway
Guan-Yu Lu1, Shu-Ting Liu2, Shih-Ming Huang2
1Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei City 114, Taiwan, ROC; Department of Pharmacy, Shalu Division, Kuang Tien General Hospital, Taichung 433, Taiwan, ROC.
Abstract:
The signaling characteristics of Na(+)/K(+)-ATPase are distinct from its ion pumping activity. Cardiac glycosides modulate the Na(+)/K(+)-ATPase protein complex upon binding, activate downstream signaling pathways and increase [Ca(2+)]i. Recent studies demonstrate that the depletion of p53 and hypoxia-induced factor 1α proteins is caused by cardiac glycosides. However, the detailed mechanisms governing this process are not well known. In this study, we showed that the depletion of p53 proteins by digoxin involved not only inhibition of protein synthesis but also inhibition at the post-transcriptional level. Post-transcriptional regulation occurs via down-regulation of SRSF3, the primary splicing factor responsible for the switch from p53α to the p53β isoform. Digoxin also modulated G2/M arrest, DNA damage and apoptosis through the p53-dependent pathway in HeLa cells. In addition, digoxin was involved in epithelial-mesenchymal-transition progression via E-cadherin reduction and snail induction. Digoxin had similar effects to caffeine, another SRSF3-reduced agent, on the cell cycle profile and DNA damage of cells. Interestingly, combined digoxin and caffeine treatment blocked cell cycle progression and conferred resistance to cell death via snail induction. These findings demonstrate that down-regulation of splicing factor, such as SRSF3, to alter cell cycle progression, cell death and invasion is a potential target for the drug repositioning of cardiac glycosides.
Insights
Cardiac glycosides like digoxin deplete p53 protein by affecting post-transcriptional regulation via SRSF3. This impacts cell cycle, DNA damage, and epithelial-mesenchymal transition, suggesting new therapeutic targets.
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Na(+)/K(+)-ATPase signaling differs from its ion pumping. Cardiac glycosides interact with this complex, activating pathways and increasing intracellular calcium.
- Cardiac glycosides are known to deplete p53 and hypoxia-induced factor 1α, but the mechanisms remain unclear.
Purpose of the Study:
- To elucidate the detailed mechanisms by which cardiac glycosides, specifically digoxin, induce p53 protein depletion.
- To investigate the role of splicing factor SRSF3 in digoxin-mediated p53 regulation.
- To explore the effects of digoxin on cell cycle, DNA damage, apoptosis, and epithelial-mesenchymal transition (EMT).
Main Methods:
- Investigated digoxin's effect on p53 protein levels, examining both synthesis and post-transcriptional regulation.
- Assessed the impact of digoxin on SRSF3 expression and its role in p53 isoform switching (p53α to p53β).
- Utilized HeLa cells to analyze digoxin's influence on G2/M arrest, DNA damage, apoptosis, E-cadherin, and snail expression.
Main Results:
- Digoxin depletes p53 not only by inhibiting synthesis but also via post-transcriptional mechanisms involving SRSF3 down-regulation.
- Digoxin treatment led to p53β isoform enrichment, G2/M arrest, DNA damage, apoptosis, and promoted EMT via reduced E-cadherin and induced snail.
- Caffeine, another SRSF3-reducing agent, showed similar effects on cell cycle and DNA damage; combined digoxin and caffeine induced snail, blocked cell cycle, and conferred resistance to cell death.
Conclusions:
- Down-regulation of splicing factor SRSF3 by digoxin alters cell cycle progression, cell death, and invasion.
- Targeting splicing factors like SRSF3 presents a potential strategy for cardiac glycoside drug repositioning.
- Digoxin's multifaceted effects on cellular processes highlight its complex pharmacology beyond ion transport.
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