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The Orphan Nuclear Receptor NR4A1 Protects Pancreatic β-Cells from Endoplasmic Reticulum (ER) Stress-mediated Apoptosis.

Yu C, Cui S, Zong C, Gao W, Xu T, Gao P, Chen J, Qin D, Guan Q, Liu Y, Fu Y, Li X, Wang X - J. Biol. Chem. (2015)

Bottom Line: This conclusion was further confirmed by experiments exploiting siRNA to knockdown NR4A1 expression in MIN6 cells or exploiting NR4A1 knock-out mice.NR4A1 overexpression in MIN6 cells or mouse islets resulted in Survivin up-regulation.In conclusion, NR4A1 protects pancreatic β-cells against ER stress-mediated apoptosis by up-regulating Survivin expression and down-regulating CHOP expression, which we termed as "positive and negative regulation."

View Article: PubMed Central - PubMed

Affiliation: From the The Department of Cell Biology, Shandong University School of Medicine, Jinan, China, 250012.

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The effect of NR4A1 overexpression on UPR.A and B, qPCR analyses of Xbp1 mRNA splicing, shown as spliced (s) relative to total (t) (A) and ATF4 expression (B) in response to 0.5 μm TG treatment of NC and OV cells at different time points. C and D, qPCR or Western blotting analysis of CHOP in NC and OV cells in response to 0.5 μm TG treatment at a series of time points. E, eif2α and phosphorylated eif2α (P-eif2α) protein expression in response to 0.5 μm TG at a series of time points. F and G, relative mRNA levels of PP1α (F) and GADD34 (G) in NC and OV cells was assessed by qPCR. H, GADD34 protein levels in NC and OV cells by Western blotting. I, the GADD34 promoter sequence contained two putative NBREs. Densitometric analyses of the Western blots are shown as curves. The data show the means of three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 versus NC.
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Figure 5: The effect of NR4A1 overexpression on UPR.A and B, qPCR analyses of Xbp1 mRNA splicing, shown as spliced (s) relative to total (t) (A) and ATF4 expression (B) in response to 0.5 μm TG treatment of NC and OV cells at different time points. C and D, qPCR or Western blotting analysis of CHOP in NC and OV cells in response to 0.5 μm TG treatment at a series of time points. E, eif2α and phosphorylated eif2α (P-eif2α) protein expression in response to 0.5 μm TG at a series of time points. F and G, relative mRNA levels of PP1α (F) and GADD34 (G) in NC and OV cells was assessed by qPCR. H, GADD34 protein levels in NC and OV cells by Western blotting. I, the GADD34 promoter sequence contained two putative NBREs. Densitometric analyses of the Western blots are shown as curves. The data show the means of three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 versus NC.

Mentions: We treated NC and OV cells with TG and harvested the cells at various time points for RNA purification or protein extraction. qPCR results showed that there was no significant difference in XBP1 splicing between NC and OV cells (Fig. 5A), but activating transcription factor 4 (ATF4) and CHOP mRNA levels were decreased in OV cells compared with NC cells (Fig. 5, B and C). CHOP protein expression also decreased in OV cells upon TG treatment compared with NC cells (Fig. 5D). Western blotting results show that phosphorylated eif2α, a subunit of eukaryotic translation initiation factor 2α, decreased substantially in OV cells compared with NC cells at 12 h and 24 h (Fig. 5E).


The Orphan Nuclear Receptor NR4A1 Protects Pancreatic β-Cells from Endoplasmic Reticulum (ER) Stress-mediated Apoptosis.

Yu C, Cui S, Zong C, Gao W, Xu T, Gao P, Chen J, Qin D, Guan Q, Liu Y, Fu Y, Li X, Wang X - J. Biol. Chem. (2015)

The effect of NR4A1 overexpression on UPR.A and B, qPCR analyses of Xbp1 mRNA splicing, shown as spliced (s) relative to total (t) (A) and ATF4 expression (B) in response to 0.5 μm TG treatment of NC and OV cells at different time points. C and D, qPCR or Western blotting analysis of CHOP in NC and OV cells in response to 0.5 μm TG treatment at a series of time points. E, eif2α and phosphorylated eif2α (P-eif2α) protein expression in response to 0.5 μm TG at a series of time points. F and G, relative mRNA levels of PP1α (F) and GADD34 (G) in NC and OV cells was assessed by qPCR. H, GADD34 protein levels in NC and OV cells by Western blotting. I, the GADD34 promoter sequence contained two putative NBREs. Densitometric analyses of the Western blots are shown as curves. The data show the means of three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 versus NC.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4543630&req=5

Figure 5: The effect of NR4A1 overexpression on UPR.A and B, qPCR analyses of Xbp1 mRNA splicing, shown as spliced (s) relative to total (t) (A) and ATF4 expression (B) in response to 0.5 μm TG treatment of NC and OV cells at different time points. C and D, qPCR or Western blotting analysis of CHOP in NC and OV cells in response to 0.5 μm TG treatment at a series of time points. E, eif2α and phosphorylated eif2α (P-eif2α) protein expression in response to 0.5 μm TG at a series of time points. F and G, relative mRNA levels of PP1α (F) and GADD34 (G) in NC and OV cells was assessed by qPCR. H, GADD34 protein levels in NC and OV cells by Western blotting. I, the GADD34 promoter sequence contained two putative NBREs. Densitometric analyses of the Western blots are shown as curves. The data show the means of three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 versus NC.
Mentions: We treated NC and OV cells with TG and harvested the cells at various time points for RNA purification or protein extraction. qPCR results showed that there was no significant difference in XBP1 splicing between NC and OV cells (Fig. 5A), but activating transcription factor 4 (ATF4) and CHOP mRNA levels were decreased in OV cells compared with NC cells (Fig. 5, B and C). CHOP protein expression also decreased in OV cells upon TG treatment compared with NC cells (Fig. 5D). Western blotting results show that phosphorylated eif2α, a subunit of eukaryotic translation initiation factor 2α, decreased substantially in OV cells compared with NC cells at 12 h and 24 h (Fig. 5E).

Bottom Line: This conclusion was further confirmed by experiments exploiting siRNA to knockdown NR4A1 expression in MIN6 cells or exploiting NR4A1 knock-out mice.NR4A1 overexpression in MIN6 cells or mouse islets resulted in Survivin up-regulation.In conclusion, NR4A1 protects pancreatic β-cells against ER stress-mediated apoptosis by up-regulating Survivin expression and down-regulating CHOP expression, which we termed as "positive and negative regulation."

View Article: PubMed Central - PubMed

Affiliation: From the The Department of Cell Biology, Shandong University School of Medicine, Jinan, China, 250012.

Show MeSH
Related in: MedlinePlus