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CArG-driven GADD45α activated by resveratrol inhibits lung cancer cells.

Shi Q, Geldenhuys W, Sutariya V, Bishayee A, Patel I, Bhatia D - Genes Cancer (2015)

Bottom Line: Therefore, induction of exogenous GADD45α resulted in growth inhibition.Since natural promoter may have antagonized effects, we tested synthetic promoter that contains either five, six or nine repeats of CArG elements essential in the Egr-1 promoter to drive the expression of GADD45α upon resveratrol treatment.Further analysis confirmed that both synthetic promoter and natural Egr-1 promoter were able to "turn on" the expression of GADD45α when combined with resveratrol, and subsequently led to suppression of cell proliferation and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Sciences, Northeast Ohio Medical University, Rootstown, Ohio, USA ; School of Biomedical Sciences, Kent State University, Kent, Ohio, USA.

ABSTRACT
We report anticarcinogenic effects of suicide gene therapy that relies on the use of resveratrol-responsive CArG elements from the Egr-1 promoter to induce GADD45α. In A549 lung cancer cells, endogenous GADD45α was not induced upon resveratrol treatment. Therefore, induction of exogenous GADD45α resulted in growth inhibition. Resveratrol transiently induced Egr-1 through ERK/JNK-ElK-1. Hence, we cloned natural or synthetic Egr-1 promoter upstream of GADD45α cDNA to create a suicide gene therapy vector. Since natural promoter may have antagonized effects, we tested synthetic promoter that contains either five, six or nine repeats of CArG elements essential in the Egr-1 promoter to drive the expression of GADD45α upon resveratrol treatment. Further analysis confirmed that both synthetic promoter and natural Egr-1 promoter were able to "turn on" the expression of GADD45α when combined with resveratrol, and subsequently led to suppression of cell proliferation and apoptosis.

No MeSH data available.


Related in: MedlinePlus

MAPK pathway is involved in resveratrol-induced Egr-1 expressionA. Cells were treated with or without 100 μM of resveratrol for 6 h after pretreatment with indicated inhibitors for 1 h, and Egr-1 protein level were examined by western blotting. B. Cells were exposed to 100 μM of resveratrol for 6 h after transfected with either Elk-1 siRNA or Luciferase siRNA. C. The summary of ERK/JNK-Elk-1-Egr-1 pathway: resveratrol sequentially activates MAPK cascade, Elk-1 and Egr-1.
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Figure 3: MAPK pathway is involved in resveratrol-induced Egr-1 expressionA. Cells were treated with or without 100 μM of resveratrol for 6 h after pretreatment with indicated inhibitors for 1 h, and Egr-1 protein level were examined by western blotting. B. Cells were exposed to 100 μM of resveratrol for 6 h after transfected with either Elk-1 siRNA or Luciferase siRNA. C. The summary of ERK/JNK-Elk-1-Egr-1 pathway: resveratrol sequentially activates MAPK cascade, Elk-1 and Egr-1.

Mentions: Activation of individual MAPK by a certain stimulus depends on the cell line, time and dose applied. Therefore, it is important to identify the involvement of each MAPK in resveratrol-mediated Egr-1 induction in A549 cells. We also tested the role of PI3K/AKT pathway as it is an important pathway to control multiple cell functions including cell proliferation, growth and survival. Cells were pretreated with p38 inhibitor (SB 203580, 10 μM), JNK inhibitor (SP 600125, 20 μM), ERK inhibitor (U-0126, 10 μM) and PI3K/AKT inhibitor (LY 294002, 10 μM) respectively for 1 h and then exposed to either 0.1% DMSO or 100 μM of resveratrol for additional 6 h. The analysis of Egr-1 protein levels indicated that SP 600125 and U-0126 were able to block resveratrol-mediated Egr-1 induction (Figure 3A, lane 6 and 8), whereas SB 203580 and LY 294002 had no significant effects on resveratrol-induced Egr-1 protein expression (Figure 3A, lane 4 and 10). Given that Elk-1 is a well-described downstream target of MAPK pathways as well as a potential regulator of Egr-1 promoter [31], we investigated whether resveratrol-induced Egr-1 expression relies on Elk-1 activity. The results demonstrated that the silencing of Elk-1 by siRNA transfection abolished the induction of Egr-1 protein by resveratrol (Figure 3B). All the data demonstrate that the normal function of JNK, ERK and Elk-1 is necessary for resveratrol-activated Egr-1 expression, and indicate that JNK/ERK-Elk-1-Egr-1 is a signaling pathway triggered by resveratrol (Figure 3C).


CArG-driven GADD45α activated by resveratrol inhibits lung cancer cells.

Shi Q, Geldenhuys W, Sutariya V, Bishayee A, Patel I, Bhatia D - Genes Cancer (2015)

MAPK pathway is involved in resveratrol-induced Egr-1 expressionA. Cells were treated with or without 100 μM of resveratrol for 6 h after pretreatment with indicated inhibitors for 1 h, and Egr-1 protein level were examined by western blotting. B. Cells were exposed to 100 μM of resveratrol for 6 h after transfected with either Elk-1 siRNA or Luciferase siRNA. C. The summary of ERK/JNK-Elk-1-Egr-1 pathway: resveratrol sequentially activates MAPK cascade, Elk-1 and Egr-1.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4482243&req=5

Figure 3: MAPK pathway is involved in resveratrol-induced Egr-1 expressionA. Cells were treated with or without 100 μM of resveratrol for 6 h after pretreatment with indicated inhibitors for 1 h, and Egr-1 protein level were examined by western blotting. B. Cells were exposed to 100 μM of resveratrol for 6 h after transfected with either Elk-1 siRNA or Luciferase siRNA. C. The summary of ERK/JNK-Elk-1-Egr-1 pathway: resveratrol sequentially activates MAPK cascade, Elk-1 and Egr-1.
Mentions: Activation of individual MAPK by a certain stimulus depends on the cell line, time and dose applied. Therefore, it is important to identify the involvement of each MAPK in resveratrol-mediated Egr-1 induction in A549 cells. We also tested the role of PI3K/AKT pathway as it is an important pathway to control multiple cell functions including cell proliferation, growth and survival. Cells were pretreated with p38 inhibitor (SB 203580, 10 μM), JNK inhibitor (SP 600125, 20 μM), ERK inhibitor (U-0126, 10 μM) and PI3K/AKT inhibitor (LY 294002, 10 μM) respectively for 1 h and then exposed to either 0.1% DMSO or 100 μM of resveratrol for additional 6 h. The analysis of Egr-1 protein levels indicated that SP 600125 and U-0126 were able to block resveratrol-mediated Egr-1 induction (Figure 3A, lane 6 and 8), whereas SB 203580 and LY 294002 had no significant effects on resveratrol-induced Egr-1 protein expression (Figure 3A, lane 4 and 10). Given that Elk-1 is a well-described downstream target of MAPK pathways as well as a potential regulator of Egr-1 promoter [31], we investigated whether resveratrol-induced Egr-1 expression relies on Elk-1 activity. The results demonstrated that the silencing of Elk-1 by siRNA transfection abolished the induction of Egr-1 protein by resveratrol (Figure 3B). All the data demonstrate that the normal function of JNK, ERK and Elk-1 is necessary for resveratrol-activated Egr-1 expression, and indicate that JNK/ERK-Elk-1-Egr-1 is a signaling pathway triggered by resveratrol (Figure 3C).

Bottom Line: Therefore, induction of exogenous GADD45α resulted in growth inhibition.Since natural promoter may have antagonized effects, we tested synthetic promoter that contains either five, six or nine repeats of CArG elements essential in the Egr-1 promoter to drive the expression of GADD45α upon resveratrol treatment.Further analysis confirmed that both synthetic promoter and natural Egr-1 promoter were able to "turn on" the expression of GADD45α when combined with resveratrol, and subsequently led to suppression of cell proliferation and apoptosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Sciences, Northeast Ohio Medical University, Rootstown, Ohio, USA ; School of Biomedical Sciences, Kent State University, Kent, Ohio, USA.

ABSTRACT
We report anticarcinogenic effects of suicide gene therapy that relies on the use of resveratrol-responsive CArG elements from the Egr-1 promoter to induce GADD45α. In A549 lung cancer cells, endogenous GADD45α was not induced upon resveratrol treatment. Therefore, induction of exogenous GADD45α resulted in growth inhibition. Resveratrol transiently induced Egr-1 through ERK/JNK-ElK-1. Hence, we cloned natural or synthetic Egr-1 promoter upstream of GADD45α cDNA to create a suicide gene therapy vector. Since natural promoter may have antagonized effects, we tested synthetic promoter that contains either five, six or nine repeats of CArG elements essential in the Egr-1 promoter to drive the expression of GADD45α upon resveratrol treatment. Further analysis confirmed that both synthetic promoter and natural Egr-1 promoter were able to "turn on" the expression of GADD45α when combined with resveratrol, and subsequently led to suppression of cell proliferation and apoptosis.

No MeSH data available.


Related in: MedlinePlus