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Methylation Status of SP1 Sites within miR-23a-27a-24-2 Promoter Region Influences Laryngeal Cancer Cell Proliferation and Apoptosis.

Wang Y, Zhang ZX, Chen S, Qiu GB, Xu ZM, Fu WN - Biomed Res Int (2016)

Bottom Line: Here we found a CG-rich region spanning two SP1 sites in the cluster promoter region.We also found that construct with two SP1 sites had highest luciferase activity and SP1 specifically bound the gene cluster promoter in vitro.We conclude that demethylated SP1 sites in miR-23a-27a-24-2 cluster upregulate the cluster expression, leading to proliferation promotion and early apoptosis inhibition in laryngeal cancer cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, China Medical University, Shenyang 110122, China.

ABSTRACT
DNA methylation plays critical roles in regulation of microRNA expression and function. miR-23a-27a-24-2 cluster has various functions and aberrant expression of the cluster is a common event in many cancers. However, whether DNA methylation influences the cluster expression and function is not reported. Here we found a CG-rich region spanning two SP1 sites in the cluster promoter region. The SP1 sites in the cluster were demethylated and methylated in Hep2 cells and HEK293 cells, respectively. Meanwhile, the cluster was significantly upregulated and downregulated in Hep2 cells and HEK293 cells, respectively. The SP1 sites were remethylated and the cluster was significantly downregulated in Hep2 cells into which methyl donor, S-adenosyl-L-methionine, was introduced. Moreover, S-adenosyl-L-methionine significantly increased Hep2 cell viability and repressed Hep2 cell early apoptosis. We also found that construct with two SP1 sites had highest luciferase activity and SP1 specifically bound the gene cluster promoter in vitro. We conclude that demethylated SP1 sites in miR-23a-27a-24-2 cluster upregulate the cluster expression, leading to proliferation promotion and early apoptosis inhibition in laryngeal cancer cells.

No MeSH data available.


Related in: MedlinePlus

Effects of DNA methylation status of miR-23a-27a-24-2 cluster promoter CG-rich region on proliferation and apoptosis. (a) Inhibition of different concentrations of SAM on Hep2 cell growth. (b) Inhibition analysis of 0.8 mmol/L SAM on Hep2 cell growth. (c) Early apoptosis analysis of different concentrations of SAM on Hep2 cells on the third day. ∗ indicates P < 0.05.
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fig2: Effects of DNA methylation status of miR-23a-27a-24-2 cluster promoter CG-rich region on proliferation and apoptosis. (a) Inhibition of different concentrations of SAM on Hep2 cell growth. (b) Inhibition analysis of 0.8 mmol/L SAM on Hep2 cell growth. (c) Early apoptosis analysis of different concentrations of SAM on Hep2 cells on the third day. ∗ indicates P < 0.05.

Mentions: To identify whether methylation status of the CpG-rich region affects Hep2 cell functions, we detected Hep2 cell proliferation and apoptosis by MTT and flow cytometry methods, respectively. MTT results showed a decreased viability tendency with the increase of concentration and treatment duration of SAM compared to the controls (Figure 2(a)). Moreover, Hep2 cell proliferation reached a peak at 0.8 mmol/L of SAM treatment on either day and showed significant differences at 0.8 mmol/L on the second and third day (Figure 2(b)). On the contrary, SAM-treated Hep2 cells displayed an increased trend in early apoptosis with an increase in SAM concentration on the third day and began to reveal a significant difference at 0.6 mmol/L compared to the controls (Figure 2(c)). We speculate that the CG-rich region hypomethylation partly regulates the Hep2 cell proliferation and early apoptosis.


Methylation Status of SP1 Sites within miR-23a-27a-24-2 Promoter Region Influences Laryngeal Cancer Cell Proliferation and Apoptosis.

Wang Y, Zhang ZX, Chen S, Qiu GB, Xu ZM, Fu WN - Biomed Res Int (2016)

Effects of DNA methylation status of miR-23a-27a-24-2 cluster promoter CG-rich region on proliferation and apoptosis. (a) Inhibition of different concentrations of SAM on Hep2 cell growth. (b) Inhibition analysis of 0.8 mmol/L SAM on Hep2 cell growth. (c) Early apoptosis analysis of different concentrations of SAM on Hep2 cells on the third day. ∗ indicates P < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4821919&req=5

fig2: Effects of DNA methylation status of miR-23a-27a-24-2 cluster promoter CG-rich region on proliferation and apoptosis. (a) Inhibition of different concentrations of SAM on Hep2 cell growth. (b) Inhibition analysis of 0.8 mmol/L SAM on Hep2 cell growth. (c) Early apoptosis analysis of different concentrations of SAM on Hep2 cells on the third day. ∗ indicates P < 0.05.
Mentions: To identify whether methylation status of the CpG-rich region affects Hep2 cell functions, we detected Hep2 cell proliferation and apoptosis by MTT and flow cytometry methods, respectively. MTT results showed a decreased viability tendency with the increase of concentration and treatment duration of SAM compared to the controls (Figure 2(a)). Moreover, Hep2 cell proliferation reached a peak at 0.8 mmol/L of SAM treatment on either day and showed significant differences at 0.8 mmol/L on the second and third day (Figure 2(b)). On the contrary, SAM-treated Hep2 cells displayed an increased trend in early apoptosis with an increase in SAM concentration on the third day and began to reveal a significant difference at 0.6 mmol/L compared to the controls (Figure 2(c)). We speculate that the CG-rich region hypomethylation partly regulates the Hep2 cell proliferation and early apoptosis.

Bottom Line: Here we found a CG-rich region spanning two SP1 sites in the cluster promoter region.We also found that construct with two SP1 sites had highest luciferase activity and SP1 specifically bound the gene cluster promoter in vitro.We conclude that demethylated SP1 sites in miR-23a-27a-24-2 cluster upregulate the cluster expression, leading to proliferation promotion and early apoptosis inhibition in laryngeal cancer cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, China Medical University, Shenyang 110122, China.

ABSTRACT
DNA methylation plays critical roles in regulation of microRNA expression and function. miR-23a-27a-24-2 cluster has various functions and aberrant expression of the cluster is a common event in many cancers. However, whether DNA methylation influences the cluster expression and function is not reported. Here we found a CG-rich region spanning two SP1 sites in the cluster promoter region. The SP1 sites in the cluster were demethylated and methylated in Hep2 cells and HEK293 cells, respectively. Meanwhile, the cluster was significantly upregulated and downregulated in Hep2 cells and HEK293 cells, respectively. The SP1 sites were remethylated and the cluster was significantly downregulated in Hep2 cells into which methyl donor, S-adenosyl-L-methionine, was introduced. Moreover, S-adenosyl-L-methionine significantly increased Hep2 cell viability and repressed Hep2 cell early apoptosis. We also found that construct with two SP1 sites had highest luciferase activity and SP1 specifically bound the gene cluster promoter in vitro. We conclude that demethylated SP1 sites in miR-23a-27a-24-2 cluster upregulate the cluster expression, leading to proliferation promotion and early apoptosis inhibition in laryngeal cancer cells.

No MeSH data available.


Related in: MedlinePlus