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Phosphatidylinositol 3-kinase signaling in proliferating cells maintains an anti-apoptotic transcriptional program mediated by inhibition of FOXO and non-canonical activation of NFkappaB transcription factors.

Terragni J, Graham JR, Adams KW, Schaffer ME, Tullai JW, Cooper GM - BMC Cell Biol. (2008)

Bottom Line: We used microarray analyses to characterize the changes in gene expression resulting from inhibition of PI 3-kinase in proliferating cells.RelB was constitutively bound to promoter regions in cells maintained in serum, however binding decreased following PI 3-kinase inhibition, indicating that PI 3-kinase signaling activates NFkappaB via the non-canonical pathway in proliferating cells.At least one-third of these genes are regulated either by FOXO transcription factors, which are activated following PI 3-kinase inhibition, or by RelB, which is activated by PI 3-kinase via the non-canonical pathway in proliferating cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Boston University, Boston MA 02215, USA. jolyon@bu.edu

ABSTRACT

Background: Phosphatidylinositol (PI) 3-kinase is activated by a variety of growth factor receptors and the PI 3-kinase/Akt signaling pathway is a key regulator of cell proliferation and survival. The downstream targets of PI 3-kinase/Akt signaling include direct regulators of cell cycle progression and apoptosis as well as a number of transcription factors. Growth factor stimulation of quiescent cells leads to robust activation of PI 3-kinase, induction of immediate-early genes, and re-entry into the cell cycle. A lower level of PI 3-kinase signaling is also required for the proliferation and survival of cells maintained in the presence of growth factors, but the gene expression program controlled by PI 3-kinase signaling in proliferating cells has not been elucidated.

Results: We used microarray analyses to characterize the changes in gene expression resulting from inhibition of PI 3-kinase in proliferating cells. The genes regulated by inhibition of PI 3-kinase in proliferating cells were distinct from genes induced by growth factor stimulation of quiescent cells and highly enriched in genes that regulate programmed cell death. Computational analyses followed by chromatin immunoprecipitations demonstrated FOXO binding to both previously known and novel sites in promoter regions of approximately one-third of the up-regulated genes, consistent with activation of FOXO1 and FOXO3a in response to inhibition of PI 3-kinase. NFkappaB binding sites were similarly identified in promoter regions of over one-third of the down-regulated genes. RelB was constitutively bound to promoter regions in cells maintained in serum, however binding decreased following PI 3-kinase inhibition, indicating that PI 3-kinase signaling activates NFkappaB via the non-canonical pathway in proliferating cells. Approximately 70% of the genes targeted by FOXO and NFkappaB regulate cell proliferation and apoptosis, including several regulators of apoptosis that were not previously known to be targeted by these transcription factors.

Conclusion: PI 3-kinase signaling in proliferating cells regulates a novel transcriptional program that is highly enriched in genes that regulate apoptosis. At least one-third of these genes are regulated either by FOXO transcription factors, which are activated following PI 3-kinase inhibition, or by RelB, which is activated by PI 3-kinase via the non-canonical pathway in proliferating cells.

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Changes in gene expression resulting from PI 3-kinase inhibition. Proliferating T98G cells in serum-containing medium were treated with 50 μM LY294002 for 2, 4 or 8 hours. Microarray analyses at each timepoint were performed on 3 independent cultures compared to untreated controls. Data are presented as the number of genes that were significantly up-regulated and down-regulated at the specified time of treatment (Log2 > 0.9, p ≤ 0.01).
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Figure 2: Changes in gene expression resulting from PI 3-kinase inhibition. Proliferating T98G cells in serum-containing medium were treated with 50 μM LY294002 for 2, 4 or 8 hours. Microarray analyses at each timepoint were performed on 3 independent cultures compared to untreated controls. Data are presented as the number of genes that were significantly up-regulated and down-regulated at the specified time of treatment (Log2 > 0.9, p ≤ 0.01).

Mentions: Microarray analyses were performed in three independent experiments to characterize the changes in gene expression resulting from PI 3-kinase inhibition. The number of genes with significantly altered expression (Log2 > 0.9; p < 0.01) after 2, 4 and 8 hours of treatment with LY294002 are shown in Fig. 2. Inhibition of PI 3-kinase for 2 hours resulted in the up-regulation of 20 genes and down-regulation of 26 genes. After 4 hours of PI 3-kinase inhibition, the number of up-regulated genes slightly increased to a total of 23, whereas the number of down-regulated genes nearly doubled, to a total of 44. The increase in the number of down-regulated genes was most likely due to the slower rates of mRNA degradation for some genes, thus requiring a longer period for significant decreases in mRNA levels to occur. After 8 hours of PI 3-kinase inhibition, the numbers of both up-regulated and down-regulated genes increased substantially to 118 and 121, respectively, possibly resulting from secondary changes in gene expression.


Phosphatidylinositol 3-kinase signaling in proliferating cells maintains an anti-apoptotic transcriptional program mediated by inhibition of FOXO and non-canonical activation of NFkappaB transcription factors.

Terragni J, Graham JR, Adams KW, Schaffer ME, Tullai JW, Cooper GM - BMC Cell Biol. (2008)

Changes in gene expression resulting from PI 3-kinase inhibition. Proliferating T98G cells in serum-containing medium were treated with 50 μM LY294002 for 2, 4 or 8 hours. Microarray analyses at each timepoint were performed on 3 independent cultures compared to untreated controls. Data are presented as the number of genes that were significantly up-regulated and down-regulated at the specified time of treatment (Log2 > 0.9, p ≤ 0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2268685&req=5

Figure 2: Changes in gene expression resulting from PI 3-kinase inhibition. Proliferating T98G cells in serum-containing medium were treated with 50 μM LY294002 for 2, 4 or 8 hours. Microarray analyses at each timepoint were performed on 3 independent cultures compared to untreated controls. Data are presented as the number of genes that were significantly up-regulated and down-regulated at the specified time of treatment (Log2 > 0.9, p ≤ 0.01).
Mentions: Microarray analyses were performed in three independent experiments to characterize the changes in gene expression resulting from PI 3-kinase inhibition. The number of genes with significantly altered expression (Log2 > 0.9; p < 0.01) after 2, 4 and 8 hours of treatment with LY294002 are shown in Fig. 2. Inhibition of PI 3-kinase for 2 hours resulted in the up-regulation of 20 genes and down-regulation of 26 genes. After 4 hours of PI 3-kinase inhibition, the number of up-regulated genes slightly increased to a total of 23, whereas the number of down-regulated genes nearly doubled, to a total of 44. The increase in the number of down-regulated genes was most likely due to the slower rates of mRNA degradation for some genes, thus requiring a longer period for significant decreases in mRNA levels to occur. After 8 hours of PI 3-kinase inhibition, the numbers of both up-regulated and down-regulated genes increased substantially to 118 and 121, respectively, possibly resulting from secondary changes in gene expression.

Bottom Line: We used microarray analyses to characterize the changes in gene expression resulting from inhibition of PI 3-kinase in proliferating cells.RelB was constitutively bound to promoter regions in cells maintained in serum, however binding decreased following PI 3-kinase inhibition, indicating that PI 3-kinase signaling activates NFkappaB via the non-canonical pathway in proliferating cells.At least one-third of these genes are regulated either by FOXO transcription factors, which are activated following PI 3-kinase inhibition, or by RelB, which is activated by PI 3-kinase via the non-canonical pathway in proliferating cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Boston University, Boston MA 02215, USA. jolyon@bu.edu

ABSTRACT

Background: Phosphatidylinositol (PI) 3-kinase is activated by a variety of growth factor receptors and the PI 3-kinase/Akt signaling pathway is a key regulator of cell proliferation and survival. The downstream targets of PI 3-kinase/Akt signaling include direct regulators of cell cycle progression and apoptosis as well as a number of transcription factors. Growth factor stimulation of quiescent cells leads to robust activation of PI 3-kinase, induction of immediate-early genes, and re-entry into the cell cycle. A lower level of PI 3-kinase signaling is also required for the proliferation and survival of cells maintained in the presence of growth factors, but the gene expression program controlled by PI 3-kinase signaling in proliferating cells has not been elucidated.

Results: We used microarray analyses to characterize the changes in gene expression resulting from inhibition of PI 3-kinase in proliferating cells. The genes regulated by inhibition of PI 3-kinase in proliferating cells were distinct from genes induced by growth factor stimulation of quiescent cells and highly enriched in genes that regulate programmed cell death. Computational analyses followed by chromatin immunoprecipitations demonstrated FOXO binding to both previously known and novel sites in promoter regions of approximately one-third of the up-regulated genes, consistent with activation of FOXO1 and FOXO3a in response to inhibition of PI 3-kinase. NFkappaB binding sites were similarly identified in promoter regions of over one-third of the down-regulated genes. RelB was constitutively bound to promoter regions in cells maintained in serum, however binding decreased following PI 3-kinase inhibition, indicating that PI 3-kinase signaling activates NFkappaB via the non-canonical pathway in proliferating cells. Approximately 70% of the genes targeted by FOXO and NFkappaB regulate cell proliferation and apoptosis, including several regulators of apoptosis that were not previously known to be targeted by these transcription factors.

Conclusion: PI 3-kinase signaling in proliferating cells regulates a novel transcriptional program that is highly enriched in genes that regulate apoptosis. At least one-third of these genes are regulated either by FOXO transcription factors, which are activated following PI 3-kinase inhibition, or by RelB, which is activated by PI 3-kinase via the non-canonical pathway in proliferating cells.

Show MeSH
Related in: MedlinePlus