Limits...
Ras oncogene-mediated progressive silencing of extracellular superoxide dismutase in tumorigenesis.

Cammarota F, de Vita G, Salvatore M, Laukkanen MO - Biomed Res Int (2015)

Bottom Line: Extracellular superoxide dismutase (SOD3) is a secreted enzyme that uses superoxide anion as a substrate in a dismutase reaction that results in the formation of hydrogen peroxide.The second class involves permanent silencing of SOD3 mediated by epigenetic DNA methylation in cells that represent more advanced cancers.Therefore, the work suggests that SOD3 belongs to the group of ras oncogene-silenced genes.

View Article: PubMed Central - PubMed

Affiliation: IRCCS SDN, 80143 Naples, Italy.

ABSTRACT
Extracellular superoxide dismutase (SOD3) is a secreted enzyme that uses superoxide anion as a substrate in a dismutase reaction that results in the formation of hydrogen peroxide. Both of these reactive oxygen species affect growth signaling in cells. Although SOD3 has growth-supporting characteristics, the expression of SOD3 is downregulated in epithelial cancer cells. In the current work, we studied the mechanisms regulating SOD3 expression in vitro using thyroid cell models representing different stages of thyroid cancer. We demonstrate that a low level of RAS activation increases SOD3 mRNA synthesis that then gradually decreases with increasing levels of RAS activation and the decreasing degree of differentiation of the cancer cells. Our data indicate that SOD3 regulation can be divided into two classes. The first class involves RAS-driven reversible regulation of SOD3 expression that can be mediated by the following mechanisms: RAS GTPase regulatory genes that are responsible for SOD3 self-regulation; RAS-stimulated p38 MAPK activation; and RAS-activated increased expression of the mir21 microRNA, which inversely correlates with sod3 mRNA expression. The second class involves permanent silencing of SOD3 mediated by epigenetic DNA methylation in cells that represent more advanced cancers. Therefore, the work suggests that SOD3 belongs to the group of ras oncogene-silenced genes.

No MeSH data available.


Related in: MedlinePlus

Effect of the demethylation reagent 5-azacytidine on SOD3 mRNA expression. (a–c) Nthy cells modeling normal human thyroid cells showed similar SOD3 mRNA expression in nontreated and treated control cells, whereas the mRNA production was significantly increased in 5-azacytidine-treated papillary TPC1 and anaplastic 8505c thyroid cancer cells. (d–f) Similar to the human cells, no alteration in sod3 mRNA was observed in rat PC Cl3 cells modeling normal rat thyroid cells after hypomethylation treatment, whereas the treatment induced increased mRNA synthesis in PTC1 and E1A oncogene-transfected cells. (g–j) The hypomethylation treatment did not affect normal rat thyroid FRLT5 cells or FRLT5 clone V13 cells harboring low 1.4-fold RAS activity, whereas a significant increase in the sod3 mRNA production was observed in clones V21 and V39 harboring 10-fold and 35-fold RAS activity, respectively. The data are expressed as the mean ± SD. The p values are represented (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4624945&req=5

fig3: Effect of the demethylation reagent 5-azacytidine on SOD3 mRNA expression. (a–c) Nthy cells modeling normal human thyroid cells showed similar SOD3 mRNA expression in nontreated and treated control cells, whereas the mRNA production was significantly increased in 5-azacytidine-treated papillary TPC1 and anaplastic 8505c thyroid cancer cells. (d–f) Similar to the human cells, no alteration in sod3 mRNA was observed in rat PC Cl3 cells modeling normal rat thyroid cells after hypomethylation treatment, whereas the treatment induced increased mRNA synthesis in PTC1 and E1A oncogene-transfected cells. (g–j) The hypomethylation treatment did not affect normal rat thyroid FRLT5 cells or FRLT5 clone V13 cells harboring low 1.4-fold RAS activity, whereas a significant increase in the sod3 mRNA production was observed in clones V21 and V39 harboring 10-fold and 35-fold RAS activity, respectively. The data are expressed as the mean ± SD. The p values are represented (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).

Mentions: The coding sequence of the sod3 gene is located in a hypermethylated CpG island [33], which suggests that methylation is involved in the regulation of the gene expression. Treatment with the hypomethylation agent 5-azacytidine did not affect SOD3 mRNA expression in human Nthy cells, rat PC Cl3 cells, rat FRLT5 cells, or FRLT5 clone V13 containing a modest 1.4-fold increased RAS activation level (Figures 3(a), 3(d), 3(g), and 3(h)); these results are consistent with our previous data showing no impact of hypomethylation on sod3 expression in aortic smooth muscle cells representing normal vascular tissue [33]. However, the 5-azacytidine treatment significantly increased SOD3 expression in the human papillary thyroid cancer TPC1 and human anaplastic thyroid cancer 8505c cells (Figures 3(b) and 3(c)). Similarly, rat thyroid PC PTC1 and PC E1A cells (Figures 3(e) and 3(f)) and FRLT5 cell clones V21 and V39 with 10-fold and 35-fold increased RAS activation, respectively, demonstrated increased sod3 mRNA expression (Figures 3(i) and 3(j)). The results thus suggest that methylation-mediated SOD3 downregulation in cancer corresponds to the degree of differentiation of the cancer and to the activation level of the RAS small GTPase.


Ras oncogene-mediated progressive silencing of extracellular superoxide dismutase in tumorigenesis.

Cammarota F, de Vita G, Salvatore M, Laukkanen MO - Biomed Res Int (2015)

Effect of the demethylation reagent 5-azacytidine on SOD3 mRNA expression. (a–c) Nthy cells modeling normal human thyroid cells showed similar SOD3 mRNA expression in nontreated and treated control cells, whereas the mRNA production was significantly increased in 5-azacytidine-treated papillary TPC1 and anaplastic 8505c thyroid cancer cells. (d–f) Similar to the human cells, no alteration in sod3 mRNA was observed in rat PC Cl3 cells modeling normal rat thyroid cells after hypomethylation treatment, whereas the treatment induced increased mRNA synthesis in PTC1 and E1A oncogene-transfected cells. (g–j) The hypomethylation treatment did not affect normal rat thyroid FRLT5 cells or FRLT5 clone V13 cells harboring low 1.4-fold RAS activity, whereas a significant increase in the sod3 mRNA production was observed in clones V21 and V39 harboring 10-fold and 35-fold RAS activity, respectively. The data are expressed as the mean ± SD. The p values are represented (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4624945&req=5

fig3: Effect of the demethylation reagent 5-azacytidine on SOD3 mRNA expression. (a–c) Nthy cells modeling normal human thyroid cells showed similar SOD3 mRNA expression in nontreated and treated control cells, whereas the mRNA production was significantly increased in 5-azacytidine-treated papillary TPC1 and anaplastic 8505c thyroid cancer cells. (d–f) Similar to the human cells, no alteration in sod3 mRNA was observed in rat PC Cl3 cells modeling normal rat thyroid cells after hypomethylation treatment, whereas the treatment induced increased mRNA synthesis in PTC1 and E1A oncogene-transfected cells. (g–j) The hypomethylation treatment did not affect normal rat thyroid FRLT5 cells or FRLT5 clone V13 cells harboring low 1.4-fold RAS activity, whereas a significant increase in the sod3 mRNA production was observed in clones V21 and V39 harboring 10-fold and 35-fold RAS activity, respectively. The data are expressed as the mean ± SD. The p values are represented (∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001).
Mentions: The coding sequence of the sod3 gene is located in a hypermethylated CpG island [33], which suggests that methylation is involved in the regulation of the gene expression. Treatment with the hypomethylation agent 5-azacytidine did not affect SOD3 mRNA expression in human Nthy cells, rat PC Cl3 cells, rat FRLT5 cells, or FRLT5 clone V13 containing a modest 1.4-fold increased RAS activation level (Figures 3(a), 3(d), 3(g), and 3(h)); these results are consistent with our previous data showing no impact of hypomethylation on sod3 expression in aortic smooth muscle cells representing normal vascular tissue [33]. However, the 5-azacytidine treatment significantly increased SOD3 expression in the human papillary thyroid cancer TPC1 and human anaplastic thyroid cancer 8505c cells (Figures 3(b) and 3(c)). Similarly, rat thyroid PC PTC1 and PC E1A cells (Figures 3(e) and 3(f)) and FRLT5 cell clones V21 and V39 with 10-fold and 35-fold increased RAS activation, respectively, demonstrated increased sod3 mRNA expression (Figures 3(i) and 3(j)). The results thus suggest that methylation-mediated SOD3 downregulation in cancer corresponds to the degree of differentiation of the cancer and to the activation level of the RAS small GTPase.

Bottom Line: Extracellular superoxide dismutase (SOD3) is a secreted enzyme that uses superoxide anion as a substrate in a dismutase reaction that results in the formation of hydrogen peroxide.The second class involves permanent silencing of SOD3 mediated by epigenetic DNA methylation in cells that represent more advanced cancers.Therefore, the work suggests that SOD3 belongs to the group of ras oncogene-silenced genes.

View Article: PubMed Central - PubMed

Affiliation: IRCCS SDN, 80143 Naples, Italy.

ABSTRACT
Extracellular superoxide dismutase (SOD3) is a secreted enzyme that uses superoxide anion as a substrate in a dismutase reaction that results in the formation of hydrogen peroxide. Both of these reactive oxygen species affect growth signaling in cells. Although SOD3 has growth-supporting characteristics, the expression of SOD3 is downregulated in epithelial cancer cells. In the current work, we studied the mechanisms regulating SOD3 expression in vitro using thyroid cell models representing different stages of thyroid cancer. We demonstrate that a low level of RAS activation increases SOD3 mRNA synthesis that then gradually decreases with increasing levels of RAS activation and the decreasing degree of differentiation of the cancer cells. Our data indicate that SOD3 regulation can be divided into two classes. The first class involves RAS-driven reversible regulation of SOD3 expression that can be mediated by the following mechanisms: RAS GTPase regulatory genes that are responsible for SOD3 self-regulation; RAS-stimulated p38 MAPK activation; and RAS-activated increased expression of the mir21 microRNA, which inversely correlates with sod3 mRNA expression. The second class involves permanent silencing of SOD3 mediated by epigenetic DNA methylation in cells that represent more advanced cancers. Therefore, the work suggests that SOD3 belongs to the group of ras oncogene-silenced genes.

No MeSH data available.


Related in: MedlinePlus