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FoxO feedback control of basal IRS-2 expression in pancreatic β-cells is distinct from that in hepatocytes.

Tsunekawa S, Demozay D, Briaud I, McCuaig J, Accili D, Stein R, Rhodes CJ - Diabetes (2011)

Bottom Line: In contrast, inhibition of phosphatidylinositol 3-kinase (PI3K) or PKB significantly increased IRS-2 levels in β-cells.ChIP assays indicated that transcription factors FoxO1 and FoxO3a associated with the IRE on the IRS-2 promoter in β-cells in a PI3K/PKB-dependent manner, whereas others, such as SREBP-1, the transcription factor binding to immunoglobulin heavy chain enhancer 3', and the aryl hydrocarbon receptor nuclear translocator (ARNT), did not.The molecular mechanism for feedback control of IRS signaling to decrease IRS-2 expression in liver and β-cells is quite distinct, with a predominant role played by FoxO3a in β-cells.

View Article: PubMed Central - PubMed

Affiliation: Kovler Diabetes Center, Department of Medicine, University of Chicago, Chicago, Illinois, USA.

ABSTRACT

Objective: Appropriate regulation of insulin receptor substrate 2 (IRS-2) expression in pancreatic β-cells is essential to adequately compensate for insulin resistance. In liver, basal IRS-2 expression is controlled via a temporal negative feedback of sterol regulatory element-binding protein 1 (SREBP-1) to antagonize transcription factors forkhead box class O (FoxO)1/FoxO3a at an insulin response element (IRE) on the IRS-2 promoter. The purpose of the study was to examine if a similar mechanism controlled IRS-2 expression in β-cells.

Research design and methods: IRS-2 mRNA and protein expression, as well as IRS-2 gene promoter activity, were examined in isolated rat islets. Specific transcription factor association with the IRE on the IRS-2 promoter was examined by chromatin immunoprecipitation (ChIP) assay, and their nuclear translocation was examined by immunofluorescence. A direct in vivo effect of insulin on control of IRS-2 expression in liver and pancreatic islets was also investigated.

Results: In IRS-2 promoter-reporter assays conducted in isolated islets, removal of the IRE decreased basal IRS-2 promoter activity in β-cells up to 80%. Activation of IRS signaling in isolated rat islets by insulin/IGF-I (used as an experimental in vitro tool) or downstream constitutive activation of protein kinase B (PKB) significantly decreased IRS-2 expression. In contrast, inhibition of phosphatidylinositol 3-kinase (PI3K) or PKB significantly increased IRS-2 levels in β-cells. ChIP assays indicated that transcription factors FoxO1 and FoxO3a associated with the IRE on the IRS-2 promoter in β-cells in a PI3K/PKB-dependent manner, whereas others, such as SREBP-1, the transcription factor binding to immunoglobulin heavy chain enhancer 3', and the aryl hydrocarbon receptor nuclear translocator (ARNT), did not. However, only FoxO3a, not FoxO1, was capable of driving IRS-2 promoter activity via the IRE in β-cells. In vivo studies showed insulin was able to suppress IRS-2 expression via activation of SREBP-1 in the liver, but this mechanism was not apparent in pancreatic islets from the same animal.

Conclusions: The molecular mechanism for feedback control of IRS signaling to decrease IRS-2 expression in liver and β-cells is quite distinct, with a predominant role played by FoxO3a in β-cells.

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Related in: MedlinePlus

FoxO3a predominately drives basal IRS-2 promoter activity β-cells. Relative FLuc activities mediated by either WT (AdV-WT-IRS-2-Luc) or IRE mutated (AdV-Mut-IRS-2-Luc) were measured as outlined in research design and methods in adenovirally infected isolated rat islets expressing GFP (AdV-GFP; control), WT FoxO1 (AdV-FoxO1-WT), or WT FoxO3a (AdV-FoxO3a-WT). Results are mean ± SEM (n ≥ 3). The * indicates statistically significant difference (P ≤ 0.05).
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Figure 5: FoxO3a predominately drives basal IRS-2 promoter activity β-cells. Relative FLuc activities mediated by either WT (AdV-WT-IRS-2-Luc) or IRE mutated (AdV-Mut-IRS-2-Luc) were measured as outlined in research design and methods in adenovirally infected isolated rat islets expressing GFP (AdV-GFP; control), WT FoxO1 (AdV-FoxO1-WT), or WT FoxO3a (AdV-FoxO3a-WT). Results are mean ± SEM (n ≥ 3). The * indicates statistically significant difference (P ≤ 0.05).

Mentions: A direct effect of FoxO1 and FoxO3a to drive IRS-2 gene transcription via the IRS-2 gene promoter IRE was examined in isolated rat islets incubated at basal 3 mmol/L glucose using the IRS-2-FLuc/TK-RLuc promoter-reporter system. In AdV-WT-IRS2-FLuc/AdV-TK-RLuc–infected rat islets additionally infected with AdV-FoxO1-WT, a modest ∼50% increase in specific WT IRS-2 promoter activity was observed above that of control islets infected with only AdV-GFP (P ≤ 0.05) (Fig. 5). In contrast, in islets additionally infected with AdV-FoxO3-WT, a significant sixfold increase in specific WT IRS-2 promoter activity was observed above that of control islets (P ≤ 0.01) (Fig. 5). In AdV-Mut-IRS2-FLuc/AdV-TK-RLuc–infected islets, where the IRE in the IRS-2 promoter-reporter vector has been mutated, the slight effect of FoxO1 to drive IRS-2 promoter activity was eliminated and the marked effect of FoxO3a inhibited ≥90% (Fig. 5). These data collectively indicate that FoxO3a drives basal IRS-2 gene transcription more effectively than FoxO1 in β-cells via the IRS-2 promoter’s IRE.


FoxO feedback control of basal IRS-2 expression in pancreatic β-cells is distinct from that in hepatocytes.

Tsunekawa S, Demozay D, Briaud I, McCuaig J, Accili D, Stein R, Rhodes CJ - Diabetes (2011)

FoxO3a predominately drives basal IRS-2 promoter activity β-cells. Relative FLuc activities mediated by either WT (AdV-WT-IRS-2-Luc) or IRE mutated (AdV-Mut-IRS-2-Luc) were measured as outlined in research design and methods in adenovirally infected isolated rat islets expressing GFP (AdV-GFP; control), WT FoxO1 (AdV-FoxO1-WT), or WT FoxO3a (AdV-FoxO3a-WT). Results are mean ± SEM (n ≥ 3). The * indicates statistically significant difference (P ≤ 0.05).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3198101&req=5

Figure 5: FoxO3a predominately drives basal IRS-2 promoter activity β-cells. Relative FLuc activities mediated by either WT (AdV-WT-IRS-2-Luc) or IRE mutated (AdV-Mut-IRS-2-Luc) were measured as outlined in research design and methods in adenovirally infected isolated rat islets expressing GFP (AdV-GFP; control), WT FoxO1 (AdV-FoxO1-WT), or WT FoxO3a (AdV-FoxO3a-WT). Results are mean ± SEM (n ≥ 3). The * indicates statistically significant difference (P ≤ 0.05).
Mentions: A direct effect of FoxO1 and FoxO3a to drive IRS-2 gene transcription via the IRS-2 gene promoter IRE was examined in isolated rat islets incubated at basal 3 mmol/L glucose using the IRS-2-FLuc/TK-RLuc promoter-reporter system. In AdV-WT-IRS2-FLuc/AdV-TK-RLuc–infected rat islets additionally infected with AdV-FoxO1-WT, a modest ∼50% increase in specific WT IRS-2 promoter activity was observed above that of control islets infected with only AdV-GFP (P ≤ 0.05) (Fig. 5). In contrast, in islets additionally infected with AdV-FoxO3-WT, a significant sixfold increase in specific WT IRS-2 promoter activity was observed above that of control islets (P ≤ 0.01) (Fig. 5). In AdV-Mut-IRS2-FLuc/AdV-TK-RLuc–infected islets, where the IRE in the IRS-2 promoter-reporter vector has been mutated, the slight effect of FoxO1 to drive IRS-2 promoter activity was eliminated and the marked effect of FoxO3a inhibited ≥90% (Fig. 5). These data collectively indicate that FoxO3a drives basal IRS-2 gene transcription more effectively than FoxO1 in β-cells via the IRS-2 promoter’s IRE.

Bottom Line: In contrast, inhibition of phosphatidylinositol 3-kinase (PI3K) or PKB significantly increased IRS-2 levels in β-cells.ChIP assays indicated that transcription factors FoxO1 and FoxO3a associated with the IRE on the IRS-2 promoter in β-cells in a PI3K/PKB-dependent manner, whereas others, such as SREBP-1, the transcription factor binding to immunoglobulin heavy chain enhancer 3', and the aryl hydrocarbon receptor nuclear translocator (ARNT), did not.The molecular mechanism for feedback control of IRS signaling to decrease IRS-2 expression in liver and β-cells is quite distinct, with a predominant role played by FoxO3a in β-cells.

View Article: PubMed Central - PubMed

Affiliation: Kovler Diabetes Center, Department of Medicine, University of Chicago, Chicago, Illinois, USA.

ABSTRACT

Objective: Appropriate regulation of insulin receptor substrate 2 (IRS-2) expression in pancreatic β-cells is essential to adequately compensate for insulin resistance. In liver, basal IRS-2 expression is controlled via a temporal negative feedback of sterol regulatory element-binding protein 1 (SREBP-1) to antagonize transcription factors forkhead box class O (FoxO)1/FoxO3a at an insulin response element (IRE) on the IRS-2 promoter. The purpose of the study was to examine if a similar mechanism controlled IRS-2 expression in β-cells.

Research design and methods: IRS-2 mRNA and protein expression, as well as IRS-2 gene promoter activity, were examined in isolated rat islets. Specific transcription factor association with the IRE on the IRS-2 promoter was examined by chromatin immunoprecipitation (ChIP) assay, and their nuclear translocation was examined by immunofluorescence. A direct in vivo effect of insulin on control of IRS-2 expression in liver and pancreatic islets was also investigated.

Results: In IRS-2 promoter-reporter assays conducted in isolated islets, removal of the IRE decreased basal IRS-2 promoter activity in β-cells up to 80%. Activation of IRS signaling in isolated rat islets by insulin/IGF-I (used as an experimental in vitro tool) or downstream constitutive activation of protein kinase B (PKB) significantly decreased IRS-2 expression. In contrast, inhibition of phosphatidylinositol 3-kinase (PI3K) or PKB significantly increased IRS-2 levels in β-cells. ChIP assays indicated that transcription factors FoxO1 and FoxO3a associated with the IRE on the IRS-2 promoter in β-cells in a PI3K/PKB-dependent manner, whereas others, such as SREBP-1, the transcription factor binding to immunoglobulin heavy chain enhancer 3', and the aryl hydrocarbon receptor nuclear translocator (ARNT), did not. However, only FoxO3a, not FoxO1, was capable of driving IRS-2 promoter activity via the IRE in β-cells. In vivo studies showed insulin was able to suppress IRS-2 expression via activation of SREBP-1 in the liver, but this mechanism was not apparent in pancreatic islets from the same animal.

Conclusions: The molecular mechanism for feedback control of IRS signaling to decrease IRS-2 expression in liver and β-cells is quite distinct, with a predominant role played by FoxO3a in β-cells.

Show MeSH
Related in: MedlinePlus