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Sulforaphane Reverses the Expression of Various Tumor Suppressor Genes by Targeting DNMT3B and HDAC1 in Human Cervical Cancer Cells.

Ali Khan M, Kedhari Sundaram M, Hamza A, Quraishi U, Gunasekera D, Ramesh L, Goala P, Al Alami U, Ansari MZ, Rizvi TA, Sharma C, Hussain A - Evid Based Complement Alternat Med (2015)

Bottom Line: Time-dependent exposure to SFN decreases the expression of DNMT3B and HDAC1 and significantly reduces the enzymatic activity of DNMTs and HDACs.Molecular modeling data suggests that SFN may interact directly with DNMT3B and HDAC1 which may explain the inhibitory action of SFN.Thus, SFN may have significant implications for epigenetic based therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Natural Science and Public Health, College of Sustainability Sciences & Humanities, Zayed University, P.O. Box 19282, Dubai, UAE.

ABSTRACT
Sulforaphane (SFN) may hinder carcinogenesis by altering epigenetic events in the cells; however, its molecular mechanisms are unclear. The present study investigates the role of SFN in modifying epigenetic events in human cervical cancer cells, HeLa. HeLa cells were treated with SFN (2.5 µM) for a period of 0, 24, 48, and 72 hours for all experiments. After treatment, expressions of DNMT3B, HDAC1, RARβ, CDH1, DAPK1, and GSTP1 were studied using RT-PCR while promoter DNA methylation of tumor suppressor genes (TSGs) was studied using MS-PCR. Inhibition assays of DNA methyl transferases (DNMTs) and histone deacetylases (HDACs) were performed at varying time points. Molecular modeling and docking studies were performed to explore the possible interaction of SFN with HDAC1 and DNMT3B. Time-dependent exposure to SFN decreases the expression of DNMT3B and HDAC1 and significantly reduces the enzymatic activity of DNMTs and HDACs. Molecular modeling data suggests that SFN may interact directly with DNMT3B and HDAC1 which may explain the inhibitory action of SFN. Interestingly, time-dependent reactivation of the studied TSGs via reversal of methylation in SFN treated cells correlates well with its impact on the epigenetic alterations accumulated during cancer development. Thus, SFN may have significant implications for epigenetic based therapy.

No MeSH data available.


Related in: MedlinePlus

Effect of SFN and TSA on HDAC1 in human cervical cancer cells (HeLa). (a) 2.5 μM of SFN and 0.05 μM TSA treatments significantly inhibit the activity of HDAC in time-dependent manner, respectively. Values are means ± SD of three independent experiments. Symbol (∗) indicates significant (P < 0.05) difference of data between control and treated cells. (b) 2.5 μM SFN treated HeLa cells show significantly time-dependent reduction in the mRNA expression of HDAC1 in comparison to untreated cells. Panel A shows β-actin expression as an internal control, Panel B shows the expression of HDAC1 on treatment with TSA, and Panel C shows the expression of HDAC1 on treatment with SFN. Lane 1 shows the expression of HDAC1 gene in untreated HeLa cells; Lanes 2, 3, and 4 show the time-dependent decrease in the expression of HDAC1 after treatment for 24, 48, and 72 h, respectively; Lane 5 shows negative control for RT-PCR.
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fig2: Effect of SFN and TSA on HDAC1 in human cervical cancer cells (HeLa). (a) 2.5 μM of SFN and 0.05 μM TSA treatments significantly inhibit the activity of HDAC in time-dependent manner, respectively. Values are means ± SD of three independent experiments. Symbol (∗) indicates significant (P < 0.05) difference of data between control and treated cells. (b) 2.5 μM SFN treated HeLa cells show significantly time-dependent reduction in the mRNA expression of HDAC1 in comparison to untreated cells. Panel A shows β-actin expression as an internal control, Panel B shows the expression of HDAC1 on treatment with TSA, and Panel C shows the expression of HDAC1 on treatment with SFN. Lane 1 shows the expression of HDAC1 gene in untreated HeLa cells; Lanes 2, 3, and 4 show the time-dependent decrease in the expression of HDAC1 after treatment for 24, 48, and 72 h, respectively; Lane 5 shows negative control for RT-PCR.

Mentions: HDACs are an enzyme family which is mainly involved in histone deacetylation and linked with the increased levels of epigenetically silenced genes in cancer. The activity of HDACs in HeLa cells was determined by treating the cells with SFN at 24, 48, and 72 h, respectively. It was observed that SFN treated HeLa cells showed a time-dependent decline of 9%, 21%, and 39% in HDACs activity (Figure 2(a)). It was also observed that the exposure of HeLa cells to HDAC inhibitor (0.05 μM TSA) showed time-dependent decrease in the activity of HDACs and caused 24% inhibition after 24 h of exposure (Figure 2(a)). Moreover, whether the decline in HDACs activity correlated with a decrease in HDAC1 expression was also investigated. It was found that exposure of HeLa cells with SFN showed a significant time-dependent decrease in the expression of HDAC1 in comparison to untreated cells (Figure 2(b)). Similar results were noticed after treatment of HeLa cells with TSA and showed significant reduced expression of HDAC1 at 24, 48, and 72 h, respectively (Figure 2(b)).


Sulforaphane Reverses the Expression of Various Tumor Suppressor Genes by Targeting DNMT3B and HDAC1 in Human Cervical Cancer Cells.

Ali Khan M, Kedhari Sundaram M, Hamza A, Quraishi U, Gunasekera D, Ramesh L, Goala P, Al Alami U, Ansari MZ, Rizvi TA, Sharma C, Hussain A - Evid Based Complement Alternat Med (2015)

Effect of SFN and TSA on HDAC1 in human cervical cancer cells (HeLa). (a) 2.5 μM of SFN and 0.05 μM TSA treatments significantly inhibit the activity of HDAC in time-dependent manner, respectively. Values are means ± SD of three independent experiments. Symbol (∗) indicates significant (P < 0.05) difference of data between control and treated cells. (b) 2.5 μM SFN treated HeLa cells show significantly time-dependent reduction in the mRNA expression of HDAC1 in comparison to untreated cells. Panel A shows β-actin expression as an internal control, Panel B shows the expression of HDAC1 on treatment with TSA, and Panel C shows the expression of HDAC1 on treatment with SFN. Lane 1 shows the expression of HDAC1 gene in untreated HeLa cells; Lanes 2, 3, and 4 show the time-dependent decrease in the expression of HDAC1 after treatment for 24, 48, and 72 h, respectively; Lane 5 shows negative control for RT-PCR.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4487331&req=5

fig2: Effect of SFN and TSA on HDAC1 in human cervical cancer cells (HeLa). (a) 2.5 μM of SFN and 0.05 μM TSA treatments significantly inhibit the activity of HDAC in time-dependent manner, respectively. Values are means ± SD of three independent experiments. Symbol (∗) indicates significant (P < 0.05) difference of data between control and treated cells. (b) 2.5 μM SFN treated HeLa cells show significantly time-dependent reduction in the mRNA expression of HDAC1 in comparison to untreated cells. Panel A shows β-actin expression as an internal control, Panel B shows the expression of HDAC1 on treatment with TSA, and Panel C shows the expression of HDAC1 on treatment with SFN. Lane 1 shows the expression of HDAC1 gene in untreated HeLa cells; Lanes 2, 3, and 4 show the time-dependent decrease in the expression of HDAC1 after treatment for 24, 48, and 72 h, respectively; Lane 5 shows negative control for RT-PCR.
Mentions: HDACs are an enzyme family which is mainly involved in histone deacetylation and linked with the increased levels of epigenetically silenced genes in cancer. The activity of HDACs in HeLa cells was determined by treating the cells with SFN at 24, 48, and 72 h, respectively. It was observed that SFN treated HeLa cells showed a time-dependent decline of 9%, 21%, and 39% in HDACs activity (Figure 2(a)). It was also observed that the exposure of HeLa cells to HDAC inhibitor (0.05 μM TSA) showed time-dependent decrease in the activity of HDACs and caused 24% inhibition after 24 h of exposure (Figure 2(a)). Moreover, whether the decline in HDACs activity correlated with a decrease in HDAC1 expression was also investigated. It was found that exposure of HeLa cells with SFN showed a significant time-dependent decrease in the expression of HDAC1 in comparison to untreated cells (Figure 2(b)). Similar results were noticed after treatment of HeLa cells with TSA and showed significant reduced expression of HDAC1 at 24, 48, and 72 h, respectively (Figure 2(b)).

Bottom Line: Time-dependent exposure to SFN decreases the expression of DNMT3B and HDAC1 and significantly reduces the enzymatic activity of DNMTs and HDACs.Molecular modeling data suggests that SFN may interact directly with DNMT3B and HDAC1 which may explain the inhibitory action of SFN.Thus, SFN may have significant implications for epigenetic based therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Natural Science and Public Health, College of Sustainability Sciences & Humanities, Zayed University, P.O. Box 19282, Dubai, UAE.

ABSTRACT
Sulforaphane (SFN) may hinder carcinogenesis by altering epigenetic events in the cells; however, its molecular mechanisms are unclear. The present study investigates the role of SFN in modifying epigenetic events in human cervical cancer cells, HeLa. HeLa cells were treated with SFN (2.5 µM) for a period of 0, 24, 48, and 72 hours for all experiments. After treatment, expressions of DNMT3B, HDAC1, RARβ, CDH1, DAPK1, and GSTP1 were studied using RT-PCR while promoter DNA methylation of tumor suppressor genes (TSGs) was studied using MS-PCR. Inhibition assays of DNA methyl transferases (DNMTs) and histone deacetylases (HDACs) were performed at varying time points. Molecular modeling and docking studies were performed to explore the possible interaction of SFN with HDAC1 and DNMT3B. Time-dependent exposure to SFN decreases the expression of DNMT3B and HDAC1 and significantly reduces the enzymatic activity of DNMTs and HDACs. Molecular modeling data suggests that SFN may interact directly with DNMT3B and HDAC1 which may explain the inhibitory action of SFN. Interestingly, time-dependent reactivation of the studied TSGs via reversal of methylation in SFN treated cells correlates well with its impact on the epigenetic alterations accumulated during cancer development. Thus, SFN may have significant implications for epigenetic based therapy.

No MeSH data available.


Related in: MedlinePlus