Limits...
IFNγ-induced suppression of β-catenin signaling: evidence for roles of Akt and 14.3.3ζ.

Nava P, Kamekura R, Quirós M, Medina-Contreras O, Hamilton RW, Kolegraff KN, Koch S, Candelario A, Romo-Parra H, Laur O, Hilgarth RS, Denning TL, Parkos CA, Nusrat A - Mol. Biol. Cell (2014)

Bottom Line: Akt1 served as a bimodal switch that promotes or inhibits β-catenin transactivation in response to IFNγ stimulation.IFNγ initially promotes β-catenin transactivation through Akt-dependent C-terminal phosphorylation of β-catenin to promote its association with 14.3.3ζ.These results outline a dual function of Akt1 that suppresses IEC proliferation during intestinal inflammation.

View Article: PubMed Central - PubMed

Affiliation: Epithelial Pathobiology and Mucosal Inflammation Research Unit, Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30322 Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Instituto Politécnico Nacional, 07360 Mexico City, Mexico.

Show MeSH

Related in: MedlinePlus

β-Catenin transactivation mediates Akt1 expression in IECs. (A) The effect of β-catenin down-regulation on Akt1 mRNA expression in response to IFNγ was analyzed by qRT-PCR. Down-regulation of β-catenin was achieved using a specific siRNA and analyzed by Western blot (inset). Experiments were performed in duplicate, and the means ± SD are shown. Akt1 mRNA levels were normalized to glyceraldehyde-3-phosphate dehydrogenase expression. *p < 0.005;**p < 0.001. (B) The effect of Akt/β-catenin signaling pathway on Akt1 mRNA expression was analyzed by qRT-PCR in control or IFNγ-treated SW480 cells. Activation of Akt/β-catenin was achieved by 14.3.3ζ overexpression or IFNγ treatment. A pharmacological inhibitor (Akt inhibitor VIII; 2.12 μM) was used to inhibit Akt. Experiments were performed in duplicate, and means ± SD are shown. *p < 0.005;**p < 0.001.
© Copyright Policy - creative-commons
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4230580&req=5

Figure 5: β-Catenin transactivation mediates Akt1 expression in IECs. (A) The effect of β-catenin down-regulation on Akt1 mRNA expression in response to IFNγ was analyzed by qRT-PCR. Down-regulation of β-catenin was achieved using a specific siRNA and analyzed by Western blot (inset). Experiments were performed in duplicate, and the means ± SD are shown. Akt1 mRNA levels were normalized to glyceraldehyde-3-phosphate dehydrogenase expression. *p < 0.005;**p < 0.001. (B) The effect of Akt/β-catenin signaling pathway on Akt1 mRNA expression was analyzed by qRT-PCR in control or IFNγ-treated SW480 cells. Activation of Akt/β-catenin was achieved by 14.3.3ζ overexpression or IFNγ treatment. A pharmacological inhibitor (Akt inhibitor VIII; 2.12 μM) was used to inhibit Akt. Experiments were performed in duplicate, and means ± SD are shown. *p < 0.005;**p < 0.001.

Mentions: Experiments were performed to investigate mechanism(s) by which IFNγ induces expression of Akt1. In colonic epithelial cells, Akt1 expression was reported to be induced by β-catenin transactivation (Dihlmann et al., 2005). Given our findings, we hypothesized that transactivation of β-catenin by Akt after IFNγ exposure at early time points (Figure 1A) was responsible for subsequent increases in Akt1protein expression. As shown in Figure 5A, IFNγ treatment for 2 h increased Akt1 mRNA expression in SW480 cells, and siRNA-mediated down-regulation of β-catenin inhibited this effect. We also investigated the possibility that β-catenin transactivation by Akt and 14.3.3ζ may regulate Akt1 expression after IFNγ treatment. Indeed, as shown in Figure 5B, IFNγ treatment resulted in an increase in Akt1 mRNA that was prevented by pharmacologic inhibition of Akt kinase activity. Furthermore, forced expression of 14.3.3ζ increased Akt1 mRNA levels that were further increased when the cells were treated with IFNγ (Figure 5B). Of importance, pharmacologic inhibition of Akt activity abrogated the increase in Akt1 mRNA mediated by 14.3.3ζ overexpression in the absence or presence of IFNγ (Figure 5B). Finally, we investigated the effect of 14.3.3ζ on the regulation of Dkk-1, a Wnt inhibitor known to be induced by IFNγ during inflammation (Nava et al., 2010). As shown in Supplemental Figure S10, up-regulation of DKK-1 mRNA levels was observed in IECs transiently expressing 14.3.3ζ. Thus, taken together, these results suggested that activation of β-catenin by Akt is necessary to induce expression of Akt1 and Dkk-1 to establish a feedback loop that inhibits Wnt/β-catenin and IEC proliferation.


IFNγ-induced suppression of β-catenin signaling: evidence for roles of Akt and 14.3.3ζ.

Nava P, Kamekura R, Quirós M, Medina-Contreras O, Hamilton RW, Kolegraff KN, Koch S, Candelario A, Romo-Parra H, Laur O, Hilgarth RS, Denning TL, Parkos CA, Nusrat A - Mol. Biol. Cell (2014)

β-Catenin transactivation mediates Akt1 expression in IECs. (A) The effect of β-catenin down-regulation on Akt1 mRNA expression in response to IFNγ was analyzed by qRT-PCR. Down-regulation of β-catenin was achieved using a specific siRNA and analyzed by Western blot (inset). Experiments were performed in duplicate, and the means ± SD are shown. Akt1 mRNA levels were normalized to glyceraldehyde-3-phosphate dehydrogenase expression. *p < 0.005;**p < 0.001. (B) The effect of Akt/β-catenin signaling pathway on Akt1 mRNA expression was analyzed by qRT-PCR in control or IFNγ-treated SW480 cells. Activation of Akt/β-catenin was achieved by 14.3.3ζ overexpression or IFNγ treatment. A pharmacological inhibitor (Akt inhibitor VIII; 2.12 μM) was used to inhibit Akt. Experiments were performed in duplicate, and means ± SD are shown. *p < 0.005;**p < 0.001.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4230580&req=5

Figure 5: β-Catenin transactivation mediates Akt1 expression in IECs. (A) The effect of β-catenin down-regulation on Akt1 mRNA expression in response to IFNγ was analyzed by qRT-PCR. Down-regulation of β-catenin was achieved using a specific siRNA and analyzed by Western blot (inset). Experiments were performed in duplicate, and the means ± SD are shown. Akt1 mRNA levels were normalized to glyceraldehyde-3-phosphate dehydrogenase expression. *p < 0.005;**p < 0.001. (B) The effect of Akt/β-catenin signaling pathway on Akt1 mRNA expression was analyzed by qRT-PCR in control or IFNγ-treated SW480 cells. Activation of Akt/β-catenin was achieved by 14.3.3ζ overexpression or IFNγ treatment. A pharmacological inhibitor (Akt inhibitor VIII; 2.12 μM) was used to inhibit Akt. Experiments were performed in duplicate, and means ± SD are shown. *p < 0.005;**p < 0.001.
Mentions: Experiments were performed to investigate mechanism(s) by which IFNγ induces expression of Akt1. In colonic epithelial cells, Akt1 expression was reported to be induced by β-catenin transactivation (Dihlmann et al., 2005). Given our findings, we hypothesized that transactivation of β-catenin by Akt after IFNγ exposure at early time points (Figure 1A) was responsible for subsequent increases in Akt1protein expression. As shown in Figure 5A, IFNγ treatment for 2 h increased Akt1 mRNA expression in SW480 cells, and siRNA-mediated down-regulation of β-catenin inhibited this effect. We also investigated the possibility that β-catenin transactivation by Akt and 14.3.3ζ may regulate Akt1 expression after IFNγ treatment. Indeed, as shown in Figure 5B, IFNγ treatment resulted in an increase in Akt1 mRNA that was prevented by pharmacologic inhibition of Akt kinase activity. Furthermore, forced expression of 14.3.3ζ increased Akt1 mRNA levels that were further increased when the cells were treated with IFNγ (Figure 5B). Of importance, pharmacologic inhibition of Akt activity abrogated the increase in Akt1 mRNA mediated by 14.3.3ζ overexpression in the absence or presence of IFNγ (Figure 5B). Finally, we investigated the effect of 14.3.3ζ on the regulation of Dkk-1, a Wnt inhibitor known to be induced by IFNγ during inflammation (Nava et al., 2010). As shown in Supplemental Figure S10, up-regulation of DKK-1 mRNA levels was observed in IECs transiently expressing 14.3.3ζ. Thus, taken together, these results suggested that activation of β-catenin by Akt is necessary to induce expression of Akt1 and Dkk-1 to establish a feedback loop that inhibits Wnt/β-catenin and IEC proliferation.

Bottom Line: Akt1 served as a bimodal switch that promotes or inhibits β-catenin transactivation in response to IFNγ stimulation.IFNγ initially promotes β-catenin transactivation through Akt-dependent C-terminal phosphorylation of β-catenin to promote its association with 14.3.3ζ.These results outline a dual function of Akt1 that suppresses IEC proliferation during intestinal inflammation.

View Article: PubMed Central - PubMed

Affiliation: Epithelial Pathobiology and Mucosal Inflammation Research Unit, Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30322 Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados, Instituto Politécnico Nacional, 07360 Mexico City, Mexico.

Show MeSH
Related in: MedlinePlus