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Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1.

Takahashi RU, Miyazaki H, Takeshita F, Yamamoto Y, Minoura K, Ono M, Kodaira M, Tamura K, Mori M, Ochiya T - Nat Commun (2015)

Bottom Line: Here, we identified microRNA-27b (miR-27b) as a key regulator for the generation of a side-population in breast cancer cells that showed CSC properties, and also found that the anti-type II diabetes (T2D) drug metformin reduced this side-population via miR-27b-mediated repression of ectonucleotide pyrophosphatase/phosphodiesterase family member 1 (ENPP1), which is involved in T2D development.ENPP1 induced the generation of the side-population via upregulation of the ABCG2 transporter.ENPP1 was also identified as a substrate of the 26S proteasome, the activity of which is downregulated in CSCs.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular and Cellular Medicine, National Cancer Center Research Institute, 1-1, Tsukiji 5-chome, Tokyo 104-0045, Japan.

ABSTRACT
Cancer stem cells (CSCs) have been identified in various types of cancer; however, the mechanisms by which cells acquire CSC properties such as drug resistance and tumour seeding ability are not fully understood. Here, we identified microRNA-27b (miR-27b) as a key regulator for the generation of a side-population in breast cancer cells that showed CSC properties, and also found that the anti-type II diabetes (T2D) drug metformin reduced this side-population via miR-27b-mediated repression of ectonucleotide pyrophosphatase/phosphodiesterase family member 1 (ENPP1), which is involved in T2D development. ENPP1 induced the generation of the side-population via upregulation of the ABCG2 transporter. ENPP1 was also identified as a substrate of the 26S proteasome, the activity of which is downregulated in CSCs. Overall, these results demonstrate that a T2D-associated gene plays an important role in tumour development and that its expression is strictly controlled at the mRNA and protein levels.

No MeSH data available.


Related in: MedlinePlus

Metformin induces miR-27b-mediated suppression of ENPP1.(a,b) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-miR-27b-DR (a) and ZR75-1-luc anti-miR-27b (b) cells incubated with metformin (0.1–10 mM) for 72 h. β-Actin was used as a loading control. (c) A schematic illustration of the miR-27b sensor construct used in the experiment shown in d. (d) Luciferase activity in MCF7 cells co-transfected with pTK-GLuc-27bs (50 ng) and pSV40-CLuc (50 ng), and then incubated with metformin for 48 h. The ratio of Gaussia to Cypridina luciferase activity (GLuc/CLuc) was determined. Data are represented as the mean±s.d. of n=3 replicates.
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f8: Metformin induces miR-27b-mediated suppression of ENPP1.(a,b) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-miR-27b-DR (a) and ZR75-1-luc anti-miR-27b (b) cells incubated with metformin (0.1–10 mM) for 72 h. β-Actin was used as a loading control. (c) A schematic illustration of the miR-27b sensor construct used in the experiment shown in d. (d) Luciferase activity in MCF7 cells co-transfected with pTK-GLuc-27bs (50 ng) and pSV40-CLuc (50 ng), and then incubated with metformin for 48 h. The ratio of Gaussia to Cypridina luciferase activity (GLuc/CLuc) was determined. Data are represented as the mean±s.d. of n=3 replicates.

Mentions: Because metformin reduces the SP fraction of breast cancer cells46 and generation of the SP fraction was induced by ENPP1, we hypothesized that metformin attenuates the chemoresistance of the SP fraction through miR-27b-mediated suppression of ENPP1. To test this hypothesis, ENPP1 expression was examined in MCF7-luc anti-miR-27b-DR and ZR75-1-luc anti-miR-27b cells that were incubated with 0.1–10 mM metformin for 72 h. Immunoblotting revealed a dose-dependent suppression of ENPP1 expression by metformin in both cell lines (Fig. 8a,b and Supplementary Fig. 12a). In addition, analysis of miR-27b expression in metformin-treated MCF7 cells using the miR-27b sensor vector (pTK-GLuc-27bs; Fig. 8c and Supplementary Fig. 2) revealed a dose-dependent suppression of luciferase activity (Fig. 8d). We also confirmed that metformin treatment reduced the SP fraction in MCF7 cells with the upregulation of miR-27b (Supplementary Fig. 12b, c). These results suggest that ENPP1 is a novel target of metformin in breast cancer cells and that metformin attenuates drug resistance through miR-27b-mediated suppression of ENPP1.


Loss of microRNA-27b contributes to breast cancer stem cell generation by activating ENPP1.

Takahashi RU, Miyazaki H, Takeshita F, Yamamoto Y, Minoura K, Ono M, Kodaira M, Tamura K, Mori M, Ochiya T - Nat Commun (2015)

Metformin induces miR-27b-mediated suppression of ENPP1.(a,b) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-miR-27b-DR (a) and ZR75-1-luc anti-miR-27b (b) cells incubated with metformin (0.1–10 mM) for 72 h. β-Actin was used as a loading control. (c) A schematic illustration of the miR-27b sensor construct used in the experiment shown in d. (d) Luciferase activity in MCF7 cells co-transfected with pTK-GLuc-27bs (50 ng) and pSV40-CLuc (50 ng), and then incubated with metformin for 48 h. The ratio of Gaussia to Cypridina luciferase activity (GLuc/CLuc) was determined. Data are represented as the mean±s.d. of n=3 replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4490376&req=5

f8: Metformin induces miR-27b-mediated suppression of ENPP1.(a,b) Immunoblot analyses of ENPP1 expression in MCF7-luc anti-miR-27b-DR (a) and ZR75-1-luc anti-miR-27b (b) cells incubated with metformin (0.1–10 mM) for 72 h. β-Actin was used as a loading control. (c) A schematic illustration of the miR-27b sensor construct used in the experiment shown in d. (d) Luciferase activity in MCF7 cells co-transfected with pTK-GLuc-27bs (50 ng) and pSV40-CLuc (50 ng), and then incubated with metformin for 48 h. The ratio of Gaussia to Cypridina luciferase activity (GLuc/CLuc) was determined. Data are represented as the mean±s.d. of n=3 replicates.
Mentions: Because metformin reduces the SP fraction of breast cancer cells46 and generation of the SP fraction was induced by ENPP1, we hypothesized that metformin attenuates the chemoresistance of the SP fraction through miR-27b-mediated suppression of ENPP1. To test this hypothesis, ENPP1 expression was examined in MCF7-luc anti-miR-27b-DR and ZR75-1-luc anti-miR-27b cells that were incubated with 0.1–10 mM metformin for 72 h. Immunoblotting revealed a dose-dependent suppression of ENPP1 expression by metformin in both cell lines (Fig. 8a,b and Supplementary Fig. 12a). In addition, analysis of miR-27b expression in metformin-treated MCF7 cells using the miR-27b sensor vector (pTK-GLuc-27bs; Fig. 8c and Supplementary Fig. 2) revealed a dose-dependent suppression of luciferase activity (Fig. 8d). We also confirmed that metformin treatment reduced the SP fraction in MCF7 cells with the upregulation of miR-27b (Supplementary Fig. 12b, c). These results suggest that ENPP1 is a novel target of metformin in breast cancer cells and that metformin attenuates drug resistance through miR-27b-mediated suppression of ENPP1.

Bottom Line: Here, we identified microRNA-27b (miR-27b) as a key regulator for the generation of a side-population in breast cancer cells that showed CSC properties, and also found that the anti-type II diabetes (T2D) drug metformin reduced this side-population via miR-27b-mediated repression of ectonucleotide pyrophosphatase/phosphodiesterase family member 1 (ENPP1), which is involved in T2D development.ENPP1 induced the generation of the side-population via upregulation of the ABCG2 transporter.ENPP1 was also identified as a substrate of the 26S proteasome, the activity of which is downregulated in CSCs.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular and Cellular Medicine, National Cancer Center Research Institute, 1-1, Tsukiji 5-chome, Tokyo 104-0045, Japan.

ABSTRACT
Cancer stem cells (CSCs) have been identified in various types of cancer; however, the mechanisms by which cells acquire CSC properties such as drug resistance and tumour seeding ability are not fully understood. Here, we identified microRNA-27b (miR-27b) as a key regulator for the generation of a side-population in breast cancer cells that showed CSC properties, and also found that the anti-type II diabetes (T2D) drug metformin reduced this side-population via miR-27b-mediated repression of ectonucleotide pyrophosphatase/phosphodiesterase family member 1 (ENPP1), which is involved in T2D development. ENPP1 induced the generation of the side-population via upregulation of the ABCG2 transporter. ENPP1 was also identified as a substrate of the 26S proteasome, the activity of which is downregulated in CSCs. Overall, these results demonstrate that a T2D-associated gene plays an important role in tumour development and that its expression is strictly controlled at the mRNA and protein levels.

No MeSH data available.


Related in: MedlinePlus