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miR-185 plays an anti-hypertrophic role in the heart via multiple targets in the calcium-signaling pathways.

Kim JO, Song DW, Kwon EJ, Hong SE, Song HK, Min CK, Kim do H - PLoS ONE (2015)

Bottom Line: The results showed that up-regulation of miR-185 led to anti-hypertrophic effects, while down-regulation led to pro-hypertrophic effects, suggesting that miR-185 has an anti-hypertrophic role in the heart.The expression of phospho-phospholamban (Thr-17), a marker of CaMKIIδ activity, was also significantly reduced by miR-185.In conclusion, miR-185 effectively blocked cardiac hypertrophy signaling through multiple targets, rendering it a potential drug target for diseases such as heart failure.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences and Systems Biology Research Center, Gwangju Institute of Science and Technology (GIST), Gwangju, Korea.

ABSTRACT
MicroRNA (miRNA) is an endogenous non-coding RNA species that either inhibits RNA translation or promotes degradation of target mRNAs. miRNAs often regulate cellular signaling by targeting multiple genes within the pathways. In the present study, using Gene Set Analysis, a useful bioinformatics tool to identify miRNAs with multiple target genes in the same pathways, we identified miR-185 as a key candidate regulator of cardiac hypertrophy. Using a mouse model, we found that miR-185 was significantly down-regulated in myocardial cells during cardiac hypertrophy induced by transverse aortic constriction. To confirm that miR-185 is an anti-hypertrophic miRNA, genetic manipulation studies such as overexpression and knock-down of miR-185 in neonatal rat ventricular myocytes were conducted. The results showed that up-regulation of miR-185 led to anti-hypertrophic effects, while down-regulation led to pro-hypertrophic effects, suggesting that miR-185 has an anti-hypertrophic role in the heart. Our study further identified Camk2d, Ncx1, and Nfatc3 as direct targets of miR-185. The activity of Nuclear Factor of Activated T-cell (NFAT) and calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ) was negatively regulated by miR-185 as assessed by NFAT-luciferase activity and western blotting. The expression of phospho-phospholamban (Thr-17), a marker of CaMKIIδ activity, was also significantly reduced by miR-185. In conclusion, miR-185 effectively blocked cardiac hypertrophy signaling through multiple targets, rendering it a potential drug target for diseases such as heart failure.

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Related in: MedlinePlus

miR-185 regulates the expression of CaMKIIδ, NCX1, and NFATC3 in cultured NRVMs.(A-C) qRT-PCR analyses of Camk2d, Ncx1 and Nfatc3 expression after transfection with miR-185 or NC. (D-F) miR-185 transfected NRVMs were lysed and analyzed by western blotting using antibodies against the proteins of interest. GAPDH or α-tubulin was used as a loading control. Representative western blots (upper) and quantified western blots (bottom). Data represent the mean ± SEM; *P < 0.05, **P < 0.001, N = 4.
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pone.0122509.g002: miR-185 regulates the expression of CaMKIIδ, NCX1, and NFATC3 in cultured NRVMs.(A-C) qRT-PCR analyses of Camk2d, Ncx1 and Nfatc3 expression after transfection with miR-185 or NC. (D-F) miR-185 transfected NRVMs were lysed and analyzed by western blotting using antibodies against the proteins of interest. GAPDH or α-tubulin was used as a loading control. Representative western blots (upper) and quantified western blots (bottom). Data represent the mean ± SEM; *P < 0.05, **P < 0.001, N = 4.

Mentions: We next screened for components in the cardiac hypertrophy signaling cascade that are controlled by miR-185. Several high score predicted targets were chosen for validation as key mediators of cardiac hypertrophy based on GSA (Figure B in S1 File). To confirm these target predictions, we transfected miR-185 into NRVMs and determined whether endogenous levels of those target genes were down-regulated. The results showed that both mRNA and protein expression levels of calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ), Na+-Ca2+ exchanger gene (NCX1/SLC8A1), and NFATC3 were significantly repressed by miR-185 compared with expression in the controls (Fig. 2A-F).


miR-185 plays an anti-hypertrophic role in the heart via multiple targets in the calcium-signaling pathways.

Kim JO, Song DW, Kwon EJ, Hong SE, Song HK, Min CK, Kim do H - PLoS ONE (2015)

miR-185 regulates the expression of CaMKIIδ, NCX1, and NFATC3 in cultured NRVMs.(A-C) qRT-PCR analyses of Camk2d, Ncx1 and Nfatc3 expression after transfection with miR-185 or NC. (D-F) miR-185 transfected NRVMs were lysed and analyzed by western blotting using antibodies against the proteins of interest. GAPDH or α-tubulin was used as a loading control. Representative western blots (upper) and quantified western blots (bottom). Data represent the mean ± SEM; *P < 0.05, **P < 0.001, N = 4.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4358957&req=5

pone.0122509.g002: miR-185 regulates the expression of CaMKIIδ, NCX1, and NFATC3 in cultured NRVMs.(A-C) qRT-PCR analyses of Camk2d, Ncx1 and Nfatc3 expression after transfection with miR-185 or NC. (D-F) miR-185 transfected NRVMs were lysed and analyzed by western blotting using antibodies against the proteins of interest. GAPDH or α-tubulin was used as a loading control. Representative western blots (upper) and quantified western blots (bottom). Data represent the mean ± SEM; *P < 0.05, **P < 0.001, N = 4.
Mentions: We next screened for components in the cardiac hypertrophy signaling cascade that are controlled by miR-185. Several high score predicted targets were chosen for validation as key mediators of cardiac hypertrophy based on GSA (Figure B in S1 File). To confirm these target predictions, we transfected miR-185 into NRVMs and determined whether endogenous levels of those target genes were down-regulated. The results showed that both mRNA and protein expression levels of calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ), Na+-Ca2+ exchanger gene (NCX1/SLC8A1), and NFATC3 were significantly repressed by miR-185 compared with expression in the controls (Fig. 2A-F).

Bottom Line: The results showed that up-regulation of miR-185 led to anti-hypertrophic effects, while down-regulation led to pro-hypertrophic effects, suggesting that miR-185 has an anti-hypertrophic role in the heart.The expression of phospho-phospholamban (Thr-17), a marker of CaMKIIδ activity, was also significantly reduced by miR-185.In conclusion, miR-185 effectively blocked cardiac hypertrophy signaling through multiple targets, rendering it a potential drug target for diseases such as heart failure.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences and Systems Biology Research Center, Gwangju Institute of Science and Technology (GIST), Gwangju, Korea.

ABSTRACT
MicroRNA (miRNA) is an endogenous non-coding RNA species that either inhibits RNA translation or promotes degradation of target mRNAs. miRNAs often regulate cellular signaling by targeting multiple genes within the pathways. In the present study, using Gene Set Analysis, a useful bioinformatics tool to identify miRNAs with multiple target genes in the same pathways, we identified miR-185 as a key candidate regulator of cardiac hypertrophy. Using a mouse model, we found that miR-185 was significantly down-regulated in myocardial cells during cardiac hypertrophy induced by transverse aortic constriction. To confirm that miR-185 is an anti-hypertrophic miRNA, genetic manipulation studies such as overexpression and knock-down of miR-185 in neonatal rat ventricular myocytes were conducted. The results showed that up-regulation of miR-185 led to anti-hypertrophic effects, while down-regulation led to pro-hypertrophic effects, suggesting that miR-185 has an anti-hypertrophic role in the heart. Our study further identified Camk2d, Ncx1, and Nfatc3 as direct targets of miR-185. The activity of Nuclear Factor of Activated T-cell (NFAT) and calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ) was negatively regulated by miR-185 as assessed by NFAT-luciferase activity and western blotting. The expression of phospho-phospholamban (Thr-17), a marker of CaMKIIδ activity, was also significantly reduced by miR-185. In conclusion, miR-185 effectively blocked cardiac hypertrophy signaling through multiple targets, rendering it a potential drug target for diseases such as heart failure.

Show MeSH
Related in: MedlinePlus