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Interleukin 6 downregulates p53 expression and activity by stimulating ribosome biogenesis: a new pathway connecting inflammation to cancer.

Brighenti E, Calabrese C, Liguori G, Giannone FA, Trerè D, Montanaro L, Derenzini M - Oncogene (2014)

Bottom Line: The p53 downregulation induced the acquisition of cellular phenotypic changes characteristic of epithelial-mesenchymal transition, such as a reduced level of E-cadherin expression, increased cell invasiveness and a decreased response to cytotoxic stresses.We found that these changes also occurred in colon epithelial cells of patients with ulcerative colitis, a very representative example of chronic inflammation at high risk for tumor development.Taken together, the present results highlight a new mechanism that may link chronic inflammation to cancer, based on p53 downregulation, which is activated by the enhancement of rRNA transcription upon IL-6 exposure.

View Article: PubMed Central - PubMed

Affiliation: Department of Experimental, Diagnostic and Specialty Medicine, Bologna University, Bologna, Italy.

ABSTRACT
Chronic inflammation is an established risk factor for the onset of cancer, and the inflammatory cytokine IL-6 has a role in tumorigenesis by enhancing proliferation and hindering apoptosis. As factors stimulating proliferation also downregulate p53 expression by enhancing ribosome biogenesis, we hypothesized that IL-6 may cause similar changes in inflamed tissues, thus activating a mechanism that favors neoplastic transformation. Here, we showed that IL-6 downregulated the expression and activity of p53 in transformed and untransformed human cell lines. This was the consequence of IL-6-dependent stimulation of c-MYC mRNA translation, which was responsible for the upregulation of rRNA transcription. The enhanced rRNA transcription stimulated the MDM2-mediated proteasomal degradation of p53, by reducing the availability of ribosome proteins for MDM2 binding. The p53 downregulation induced the acquisition of cellular phenotypic changes characteristic of epithelial-mesenchymal transition, such as a reduced level of E-cadherin expression, increased cell invasiveness and a decreased response to cytotoxic stresses. We found that these changes also occurred in colon epithelial cells of patients with ulcerative colitis, a very representative example of chronic inflammation at high risk for tumor development. Histochemical and immunohistochemical analysis of colon biopsy samples showed an upregulation of ribosome biogenesis, a reduced expression of p53, together with a focal reduction or absence of E-cadherin expression in chronic colitis in comparison with normal mucosa samples. These changes disappeared after treatment with anti-inflammatory drugs. Taken together, the present results highlight a new mechanism that may link chronic inflammation to cancer, based on p53 downregulation, which is activated by the enhancement of rRNA transcription upon IL-6 exposure.

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Related in: MedlinePlus

IL-6 induces EMT in a p53-dependent manner. (a) Representative western blot analysis of E-cadherin and SLUG expression in NCM460, HepG2 and HCT116 p53−/− cells exposed to IL-6 for 24 h. NCM460 were either (p53−) or not (SCR) silenced for TP53 expression. (b) Visualization of SLUG and E-cadherin distribution in control and IL-6-treated NCM460 cells. Cells were labeled with monoclonal antibodies versus SLUG or E-cadherin; the antibodies were revealed by FITC-conjugated secondary antibodies. Nuclei were stained with 4',6-diamidino-2-phenylindole (DAPI). Scale bar=40 μm. (c) Invasion assay of control (SCR) and TP53-silenced (p53−) NCM460 and HepG2 cells. The cells were exposed, 48 h after the end of the silencing procedure, to IL-6 for 24 h. (d) Real-time–PCR evaluation of 45S rRNA and western blot analysis of p53 expression in NCM460 and HepG2 cells transfected with control sequences (SCR) and in POLR1A-silenced cells (Pol1−). At 48 h after the end of the silencing procedure cells were exposed to IL-6 for 24 h. (e) Western blot and densitometric analysis of E-cadherin expression in control (SCR) and POLR1A-silenced (Pol1−) HepG2 cells. At 48 h after the end of the silencing procedure the cells were exposed to IL-6 for 24 h. (f) Invasion assay of control (SCR) and POLR1A-silenced NCM460 and HepG2 cells. At 48 h after the end of the silencing procedure, the cells were exposed to IL-6 for 24 h. Histograms show the values (mean±s.d.) of three experiments. *P<0.05; **P<0.01; n.s., not significant.
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fig3: IL-6 induces EMT in a p53-dependent manner. (a) Representative western blot analysis of E-cadherin and SLUG expression in NCM460, HepG2 and HCT116 p53−/− cells exposed to IL-6 for 24 h. NCM460 were either (p53−) or not (SCR) silenced for TP53 expression. (b) Visualization of SLUG and E-cadherin distribution in control and IL-6-treated NCM460 cells. Cells were labeled with monoclonal antibodies versus SLUG or E-cadherin; the antibodies were revealed by FITC-conjugated secondary antibodies. Nuclei were stained with 4',6-diamidino-2-phenylindole (DAPI). Scale bar=40 μm. (c) Invasion assay of control (SCR) and TP53-silenced (p53−) NCM460 and HepG2 cells. The cells were exposed, 48 h after the end of the silencing procedure, to IL-6 for 24 h. (d) Real-time–PCR evaluation of 45S rRNA and western blot analysis of p53 expression in NCM460 and HepG2 cells transfected with control sequences (SCR) and in POLR1A-silenced cells (Pol1−). At 48 h after the end of the silencing procedure cells were exposed to IL-6 for 24 h. (e) Western blot and densitometric analysis of E-cadherin expression in control (SCR) and POLR1A-silenced (Pol1−) HepG2 cells. At 48 h after the end of the silencing procedure the cells were exposed to IL-6 for 24 h. (f) Invasion assay of control (SCR) and POLR1A-silenced NCM460 and HepG2 cells. At 48 h after the end of the silencing procedure, the cells were exposed to IL-6 for 24 h. Histograms show the values (mean±s.d.) of three experiments. *P<0.05; **P<0.01; n.s., not significant.

Mentions: Downregulation of E-cadherin expression occurs very frequently during the progression of malignant epithelial tumors,23 and the loss of E-cadherin expression is responsible for the loss of intercellular adhesion during invasion.24 The E-cadherin expression is repressed by the transcription factor SLUG, responsible for EMT.25 As p53 controls the MDM2-mediated degradation of SLUG, 26 we wondered whether IL-6 could induce a reduction of E-cadherin expression by increasing the level of SLUG in a p53-dependent manner. For this purpose, we analyzed E-cadherin and SLUG protein expression: we found that at 24 h IL-6 stimulation increased the level of SLUG and reduced the expression of E-cadherin in NCM460 and HepG2 cell lines (Figure 3a). We did not observe any significant variation in the expression of E-cadherin and SLUG in TP53-silenced NCM460 cells (see the efficacy of TP53 silencing in Supplementary Figure 5).On the other hand, to avoid any misinterpretation on the role of p53 in reducing the level of E-cadherin in IL-6-treated cells, we used the HCT116 p53−/− cell line and analyzed the level of both E-cadherin and SLUG after IL-6 treatment. As shown in Figure 3a, IL-6 treatment did not modify the amount of both these proteins in HCT116 p53−/− cells, thus indicating the crucial role of p53 in this mechanism.


Interleukin 6 downregulates p53 expression and activity by stimulating ribosome biogenesis: a new pathway connecting inflammation to cancer.

Brighenti E, Calabrese C, Liguori G, Giannone FA, Trerè D, Montanaro L, Derenzini M - Oncogene (2014)

IL-6 induces EMT in a p53-dependent manner. (a) Representative western blot analysis of E-cadherin and SLUG expression in NCM460, HepG2 and HCT116 p53−/− cells exposed to IL-6 for 24 h. NCM460 were either (p53−) or not (SCR) silenced for TP53 expression. (b) Visualization of SLUG and E-cadherin distribution in control and IL-6-treated NCM460 cells. Cells were labeled with monoclonal antibodies versus SLUG or E-cadherin; the antibodies were revealed by FITC-conjugated secondary antibodies. Nuclei were stained with 4',6-diamidino-2-phenylindole (DAPI). Scale bar=40 μm. (c) Invasion assay of control (SCR) and TP53-silenced (p53−) NCM460 and HepG2 cells. The cells were exposed, 48 h after the end of the silencing procedure, to IL-6 for 24 h. (d) Real-time–PCR evaluation of 45S rRNA and western blot analysis of p53 expression in NCM460 and HepG2 cells transfected with control sequences (SCR) and in POLR1A-silenced cells (Pol1−). At 48 h after the end of the silencing procedure cells were exposed to IL-6 for 24 h. (e) Western blot and densitometric analysis of E-cadherin expression in control (SCR) and POLR1A-silenced (Pol1−) HepG2 cells. At 48 h after the end of the silencing procedure the cells were exposed to IL-6 for 24 h. (f) Invasion assay of control (SCR) and POLR1A-silenced NCM460 and HepG2 cells. At 48 h after the end of the silencing procedure, the cells were exposed to IL-6 for 24 h. Histograms show the values (mean±s.d.) of three experiments. *P<0.05; **P<0.01; n.s., not significant.
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Related In: Results  -  Collection

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fig3: IL-6 induces EMT in a p53-dependent manner. (a) Representative western blot analysis of E-cadherin and SLUG expression in NCM460, HepG2 and HCT116 p53−/− cells exposed to IL-6 for 24 h. NCM460 were either (p53−) or not (SCR) silenced for TP53 expression. (b) Visualization of SLUG and E-cadherin distribution in control and IL-6-treated NCM460 cells. Cells were labeled with monoclonal antibodies versus SLUG or E-cadherin; the antibodies were revealed by FITC-conjugated secondary antibodies. Nuclei were stained with 4',6-diamidino-2-phenylindole (DAPI). Scale bar=40 μm. (c) Invasion assay of control (SCR) and TP53-silenced (p53−) NCM460 and HepG2 cells. The cells were exposed, 48 h after the end of the silencing procedure, to IL-6 for 24 h. (d) Real-time–PCR evaluation of 45S rRNA and western blot analysis of p53 expression in NCM460 and HepG2 cells transfected with control sequences (SCR) and in POLR1A-silenced cells (Pol1−). At 48 h after the end of the silencing procedure cells were exposed to IL-6 for 24 h. (e) Western blot and densitometric analysis of E-cadherin expression in control (SCR) and POLR1A-silenced (Pol1−) HepG2 cells. At 48 h after the end of the silencing procedure the cells were exposed to IL-6 for 24 h. (f) Invasion assay of control (SCR) and POLR1A-silenced NCM460 and HepG2 cells. At 48 h after the end of the silencing procedure, the cells were exposed to IL-6 for 24 h. Histograms show the values (mean±s.d.) of three experiments. *P<0.05; **P<0.01; n.s., not significant.
Mentions: Downregulation of E-cadherin expression occurs very frequently during the progression of malignant epithelial tumors,23 and the loss of E-cadherin expression is responsible for the loss of intercellular adhesion during invasion.24 The E-cadherin expression is repressed by the transcription factor SLUG, responsible for EMT.25 As p53 controls the MDM2-mediated degradation of SLUG, 26 we wondered whether IL-6 could induce a reduction of E-cadherin expression by increasing the level of SLUG in a p53-dependent manner. For this purpose, we analyzed E-cadherin and SLUG protein expression: we found that at 24 h IL-6 stimulation increased the level of SLUG and reduced the expression of E-cadherin in NCM460 and HepG2 cell lines (Figure 3a). We did not observe any significant variation in the expression of E-cadherin and SLUG in TP53-silenced NCM460 cells (see the efficacy of TP53 silencing in Supplementary Figure 5).On the other hand, to avoid any misinterpretation on the role of p53 in reducing the level of E-cadherin in IL-6-treated cells, we used the HCT116 p53−/− cell line and analyzed the level of both E-cadherin and SLUG after IL-6 treatment. As shown in Figure 3a, IL-6 treatment did not modify the amount of both these proteins in HCT116 p53−/− cells, thus indicating the crucial role of p53 in this mechanism.

Bottom Line: The p53 downregulation induced the acquisition of cellular phenotypic changes characteristic of epithelial-mesenchymal transition, such as a reduced level of E-cadherin expression, increased cell invasiveness and a decreased response to cytotoxic stresses.We found that these changes also occurred in colon epithelial cells of patients with ulcerative colitis, a very representative example of chronic inflammation at high risk for tumor development.Taken together, the present results highlight a new mechanism that may link chronic inflammation to cancer, based on p53 downregulation, which is activated by the enhancement of rRNA transcription upon IL-6 exposure.

View Article: PubMed Central - PubMed

Affiliation: Department of Experimental, Diagnostic and Specialty Medicine, Bologna University, Bologna, Italy.

ABSTRACT
Chronic inflammation is an established risk factor for the onset of cancer, and the inflammatory cytokine IL-6 has a role in tumorigenesis by enhancing proliferation and hindering apoptosis. As factors stimulating proliferation also downregulate p53 expression by enhancing ribosome biogenesis, we hypothesized that IL-6 may cause similar changes in inflamed tissues, thus activating a mechanism that favors neoplastic transformation. Here, we showed that IL-6 downregulated the expression and activity of p53 in transformed and untransformed human cell lines. This was the consequence of IL-6-dependent stimulation of c-MYC mRNA translation, which was responsible for the upregulation of rRNA transcription. The enhanced rRNA transcription stimulated the MDM2-mediated proteasomal degradation of p53, by reducing the availability of ribosome proteins for MDM2 binding. The p53 downregulation induced the acquisition of cellular phenotypic changes characteristic of epithelial-mesenchymal transition, such as a reduced level of E-cadherin expression, increased cell invasiveness and a decreased response to cytotoxic stresses. We found that these changes also occurred in colon epithelial cells of patients with ulcerative colitis, a very representative example of chronic inflammation at high risk for tumor development. Histochemical and immunohistochemical analysis of colon biopsy samples showed an upregulation of ribosome biogenesis, a reduced expression of p53, together with a focal reduction or absence of E-cadherin expression in chronic colitis in comparison with normal mucosa samples. These changes disappeared after treatment with anti-inflammatory drugs. Taken together, the present results highlight a new mechanism that may link chronic inflammation to cancer, based on p53 downregulation, which is activated by the enhancement of rRNA transcription upon IL-6 exposure.

Show MeSH
Related in: MedlinePlus