Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway

Bruna Corominas-Faja, Sílvia Cufí, Cristina Oliveras-Ferraros, Elisabet Cuyàs, Eugeni López-Bonet, Ruth Lupu, Tomás Alarcón, Luciano Vellon, Juan Manuel Iglesias, Olatz Leis, Ángel G. Martín, Alejandro Vazquez-Martin, Javier A. Menendez

Research output: Contribution to journalArticle

63 Citations (Scopus)

Abstract

Energy metabolism plasticity enables stemness programs during the reprogramming of somatic cells to an induced pluripotent stem cell (iPSC) state. This relationship may introduce a new era in the understanding of Warburg's theory on the metabolic origin of cancer at the level of cancer stem cells (CSCs). Here, we used Yamanaka's stem cell technology in an attempt to create stable CSC research lines in which to dissect the transcriptional control of mTO R - the master switch of cellular catabolism and anabolism - in CSC-like states. The rare colonies with iPSC-like morphology, obtained following the viral transduction of the Oct4, Sox2, Klf4, and c-Myc (OSKM) stemness factors into MCF-7 luminal-like breast cancer cells (MCF-7/Rep), demonstrated an intermediate state between cancer cells and bona fide iPSCs. MCF-7/Rep cells notably overexpressed SOX2 and stage-specific embryonic antigen (SSEA)-4 proteins; however, other stemness-related markers (OCT4, NANOG, SSEA-1, TRA-1-60, and TRA-1-81) were found at low to moderate levels. The transcriptional analyses of OSKM factors confirmed the strong but unique reactivation of the endogenous Sox2 stemness gene accompanied by the silencing of the exogenous Sox2 transgene in MCF-7/Rep cells. Some but not all MCF-7/Rep cells acquired strong alkaline phosphatase (AP) activity compared with MCF-7 parental cells. SOX2-overexpressing MCF-7/Rep cells contained drastically higher percentages of CD44+ and ALDEFLUOR-stained ALDHbright cells than MCF-7 parental cells. The overlap between differentially expressed mTO R signaling-related genes in 3 different SOX2-overexpressing CSC-like cell lines revealed a notable downregulation of 3 genes, PRKAA1 (which codes for the catalytic α 1 subunit of AMPK), DDIT4/REDD1 (a stress response gene that operates as a negative regulator of mTO R), and DEPTOR (a naturally occurring endogenous inhibitor of mTO R activity). The insulin-receptor gene (INSR) was differentially upregulated in MCF-7/Rep cells. Consistent with the downregulation of AMPK expression, immunoblotting procedures confirmed upregulation of p70S6K and increased phosphorylation of mTO R in Sox2-overexpressing CSC-like cell populations. Using an in vitro model of the de novo generation of CSC-like states through the nuclear reprogramming of an established breast cancer cell line, we reveal that the transcriptional suppression of mTO R repressors is an intrinsic process occurring during the acquisition of CSC-like properties by differentiated populations of luminal-like breast cancer cells. This approach may provide a new path for obtaining information about preventing the appearance of CSCs through the modulation of the AMPK/mTO R pathway.

Original languageEnglish (US)
Pages (from-to)3109-3124
Number of pages16
JournalCell Cycle
Volume12
Issue number18
DOIs
StatePublished - Sep 15 2013

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Neoplastic Stem Cells
MCF-7 Cells
Transcriptional Activation
Breast Neoplasms
AMP-Activated Protein Kinases
Neoplasms
Induced Pluripotent Stem Cells
Cell Line
Genes
Down-Regulation
CD15 Antigens
70-kDa Ribosomal Protein S6 Kinases
Stem Cell Research
Cellular Reprogramming
Insulin Receptor
Gene Silencing
Transgenes
Immunoblotting
Energy Metabolism
Population

Keywords

  • AMPK
  • Breast cancer
  • Cancer stem cells
  • MTOR
  • Reprogramming
  • SOX2

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology
  • Developmental Biology

Cite this

Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway. / Corominas-Faja, Bruna; Cufí, Sílvia; Oliveras-Ferraros, Cristina; Cuyàs, Elisabet; López-Bonet, Eugeni; Lupu, Ruth; Alarcón, Tomás; Vellon, Luciano; Iglesias, Juan Manuel; Leis, Olatz; Martín, Ángel G.; Vazquez-Martin, Alejandro; Menendez, Javier A.

In: Cell Cycle, Vol. 12, No. 18, 15.09.2013, p. 3109-3124.

Research output: Contribution to journalArticle

Corominas-Faja, B, Cufí, S, Oliveras-Ferraros, C, Cuyàs, E, López-Bonet, E, Lupu, R, Alarcón, T, Vellon, L, Iglesias, JM, Leis, O, Martín, ÁG, Vazquez-Martin, A & Menendez, JA 2013, 'Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway', Cell Cycle, vol. 12, no. 18, pp. 3109-3124. https://doi.org/10.4161/cc.26173
Corominas-Faja, Bruna ; Cufí, Sílvia ; Oliveras-Ferraros, Cristina ; Cuyàs, Elisabet ; López-Bonet, Eugeni ; Lupu, Ruth ; Alarcón, Tomás ; Vellon, Luciano ; Iglesias, Juan Manuel ; Leis, Olatz ; Martín, Ángel G. ; Vazquez-Martin, Alejandro ; Menendez, Javier A. / Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway. In: Cell Cycle. 2013 ; Vol. 12, No. 18. pp. 3109-3124.
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T1 - Nuclear reprogramming of luminal-like breast cancer cells generates Sox2-overexpressing cancer stem-like cellular states harboring transcriptional activation of the mTOR pathway

AU - Corominas-Faja, Bruna

AU - Cufí, Sílvia

AU - Oliveras-Ferraros, Cristina

AU - Cuyàs, Elisabet

AU - López-Bonet, Eugeni

AU - Lupu, Ruth

AU - Alarcón, Tomás

AU - Vellon, Luciano

AU - Iglesias, Juan Manuel

AU - Leis, Olatz

AU - Martín, Ángel G.

AU - Vazquez-Martin, Alejandro

AU - Menendez, Javier A.

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N2 - Energy metabolism plasticity enables stemness programs during the reprogramming of somatic cells to an induced pluripotent stem cell (iPSC) state. This relationship may introduce a new era in the understanding of Warburg's theory on the metabolic origin of cancer at the level of cancer stem cells (CSCs). Here, we used Yamanaka's stem cell technology in an attempt to create stable CSC research lines in which to dissect the transcriptional control of mTO R - the master switch of cellular catabolism and anabolism - in CSC-like states. The rare colonies with iPSC-like morphology, obtained following the viral transduction of the Oct4, Sox2, Klf4, and c-Myc (OSKM) stemness factors into MCF-7 luminal-like breast cancer cells (MCF-7/Rep), demonstrated an intermediate state between cancer cells and bona fide iPSCs. MCF-7/Rep cells notably overexpressed SOX2 and stage-specific embryonic antigen (SSEA)-4 proteins; however, other stemness-related markers (OCT4, NANOG, SSEA-1, TRA-1-60, and TRA-1-81) were found at low to moderate levels. The transcriptional analyses of OSKM factors confirmed the strong but unique reactivation of the endogenous Sox2 stemness gene accompanied by the silencing of the exogenous Sox2 transgene in MCF-7/Rep cells. Some but not all MCF-7/Rep cells acquired strong alkaline phosphatase (AP) activity compared with MCF-7 parental cells. SOX2-overexpressing MCF-7/Rep cells contained drastically higher percentages of CD44+ and ALDEFLUOR-stained ALDHbright cells than MCF-7 parental cells. The overlap between differentially expressed mTO R signaling-related genes in 3 different SOX2-overexpressing CSC-like cell lines revealed a notable downregulation of 3 genes, PRKAA1 (which codes for the catalytic α 1 subunit of AMPK), DDIT4/REDD1 (a stress response gene that operates as a negative regulator of mTO R), and DEPTOR (a naturally occurring endogenous inhibitor of mTO R activity). The insulin-receptor gene (INSR) was differentially upregulated in MCF-7/Rep cells. Consistent with the downregulation of AMPK expression, immunoblotting procedures confirmed upregulation of p70S6K and increased phosphorylation of mTO R in Sox2-overexpressing CSC-like cell populations. Using an in vitro model of the de novo generation of CSC-like states through the nuclear reprogramming of an established breast cancer cell line, we reveal that the transcriptional suppression of mTO R repressors is an intrinsic process occurring during the acquisition of CSC-like properties by differentiated populations of luminal-like breast cancer cells. This approach may provide a new path for obtaining information about preventing the appearance of CSCs through the modulation of the AMPK/mTO R pathway.

AB - Energy metabolism plasticity enables stemness programs during the reprogramming of somatic cells to an induced pluripotent stem cell (iPSC) state. This relationship may introduce a new era in the understanding of Warburg's theory on the metabolic origin of cancer at the level of cancer stem cells (CSCs). Here, we used Yamanaka's stem cell technology in an attempt to create stable CSC research lines in which to dissect the transcriptional control of mTO R - the master switch of cellular catabolism and anabolism - in CSC-like states. The rare colonies with iPSC-like morphology, obtained following the viral transduction of the Oct4, Sox2, Klf4, and c-Myc (OSKM) stemness factors into MCF-7 luminal-like breast cancer cells (MCF-7/Rep), demonstrated an intermediate state between cancer cells and bona fide iPSCs. MCF-7/Rep cells notably overexpressed SOX2 and stage-specific embryonic antigen (SSEA)-4 proteins; however, other stemness-related markers (OCT4, NANOG, SSEA-1, TRA-1-60, and TRA-1-81) were found at low to moderate levels. The transcriptional analyses of OSKM factors confirmed the strong but unique reactivation of the endogenous Sox2 stemness gene accompanied by the silencing of the exogenous Sox2 transgene in MCF-7/Rep cells. Some but not all MCF-7/Rep cells acquired strong alkaline phosphatase (AP) activity compared with MCF-7 parental cells. SOX2-overexpressing MCF-7/Rep cells contained drastically higher percentages of CD44+ and ALDEFLUOR-stained ALDHbright cells than MCF-7 parental cells. The overlap between differentially expressed mTO R signaling-related genes in 3 different SOX2-overexpressing CSC-like cell lines revealed a notable downregulation of 3 genes, PRKAA1 (which codes for the catalytic α 1 subunit of AMPK), DDIT4/REDD1 (a stress response gene that operates as a negative regulator of mTO R), and DEPTOR (a naturally occurring endogenous inhibitor of mTO R activity). The insulin-receptor gene (INSR) was differentially upregulated in MCF-7/Rep cells. Consistent with the downregulation of AMPK expression, immunoblotting procedures confirmed upregulation of p70S6K and increased phosphorylation of mTO R in Sox2-overexpressing CSC-like cell populations. Using an in vitro model of the de novo generation of CSC-like states through the nuclear reprogramming of an established breast cancer cell line, we reveal that the transcriptional suppression of mTO R repressors is an intrinsic process occurring during the acquisition of CSC-like properties by differentiated populations of luminal-like breast cancer cells. This approach may provide a new path for obtaining information about preventing the appearance of CSCs through the modulation of the AMPK/mTO R pathway.

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KW - Cancer stem cells

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