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Cdh2 stabilizes FGFR1 and contributes to primed-state pluripotency in mouse epiblast stem cells.

Takehara T, Teramura T, Onodera Y, Frampton J, Fukuda K - Sci Rep (2015)

Bottom Line: Western blot analysis and co-immunoprecipitation assays revealed that Cdh2 stabilizes FGFR1 in mEpiSCs.Furthermore, stable transfection of mESCs with Cdh2 cDNA followed by FGF2 supplementation accelerated cell differentiation.Thus, Cdh2 contributes to the establishment and maintenance of FGF signaling-dependent self-renewal in mEpiSCs through stabilization of FGFR1.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell Biology for Regenerative Medicine, Institute of Advanced Clinical Medicine, Kindai University Faculty of Medicine, 377-2 Ohnohigashi, Osaka-sayama, Osaka, Japan 5898511.

ABSTRACT
The cell adhesion molecule Cadherin 2 (Cdh2) plays important roles in somatic cell adhesion, proliferation and migration. Cdh2 is also highly expressed in mouse epiblast stem cells (mEpiSCs), but its function in these cells is unknown. To understand the function of Cdh2 in mEpiSCs, we compared the expression of pluripotency-related genes in mEpiSCs and mouse embryonic stem cells (mESCs) after either Cdh2 knockdown or Cdh2 over-expression. Introduction of specific siRNA against Cdh2 led to attenuation of pluripotency-related genes. Pluripotent gene expression was not recovered by over-expression of Cdh1 following Cdh2 knockdown. Western blot analysis and co-immunoprecipitation assays revealed that Cdh2 stabilizes FGFR1 in mEpiSCs. Furthermore, stable transfection of mESCs with Cdh2 cDNA followed by FGF2 supplementation accelerated cell differentiation. Thus, Cdh2 contributes to the establishment and maintenance of FGF signaling-dependent self-renewal in mEpiSCs through stabilization of FGFR1.

No MeSH data available.


Related in: MedlinePlus

Cdh2 is the major cadherin-type expressed by mEpiSCs.(A) qRT-PCR analysis for classical and atypical cadherins in mESCs and mEpiSCs. Bars represent the mean values of triplicates. White bars indicate gene expression in the mESCs and gray bars indicate gene expression in the mEpiSCs. (B) WB analysis for Cdh1 and Cdh2 in mESCs and mEpiSCs. (C) Immunofluorescent staining for Cdh1 and Cdh2 in mESCs and mEpiSCs. The scale bars are 100 μm.
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f1: Cdh2 is the major cadherin-type expressed by mEpiSCs.(A) qRT-PCR analysis for classical and atypical cadherins in mESCs and mEpiSCs. Bars represent the mean values of triplicates. White bars indicate gene expression in the mESCs and gray bars indicate gene expression in the mEpiSCs. (B) WB analysis for Cdh1 and Cdh2 in mESCs and mEpiSCs. (C) Immunofluorescent staining for Cdh1 and Cdh2 in mESCs and mEpiSCs. The scale bars are 100 μm.

Mentions: We first analyzed the expression of a variety of classical and atypical cadherin genes: Cdh1 (Epithelial-cadherin), Cdh2 (Neuronal-cadherin), Cdh3 (Placental-cadherin), Cdh4 (Retinal-cadherin), Cdh5 (Vascular Endothelial-cadherin), Cdh12 (Neuronal-cadherin II), Cdh13 (T-cadherin, heart-cadherin) and Cdh15 (Myotubular-cadherin) by quantitative RT-PCR (qRT-PCR). In mESCs, Cdh1 was the most highly expressed of the cadherin genes, although Cdh3 was also expressed. In contrast, mEpiSCs predominantly expressed Cdh1 and Cdh2 (Fig. 1A). We next determined Cdh1 and Cdh2 protein expression in mESCs and mEpiSCs by Western blot (WB) analysis (Fig. 1B). Cdh1 was abundant in mESCs, with only low levels of Cdh2. In contrast, Cdh2 was abundant in mEpiSCs, with almost no Cdh1. Immunofluorescence confirmed the expression of Cdh1 and Cdh2 in the mESCs and mEpiSCs, respectively (Fig. 1C). From these results, we conclude that Cdh1 is a mESC status-specific cadherin and that Cdh2 is a mEpiSC status-specific cadherin.


Cdh2 stabilizes FGFR1 and contributes to primed-state pluripotency in mouse epiblast stem cells.

Takehara T, Teramura T, Onodera Y, Frampton J, Fukuda K - Sci Rep (2015)

Cdh2 is the major cadherin-type expressed by mEpiSCs.(A) qRT-PCR analysis for classical and atypical cadherins in mESCs and mEpiSCs. Bars represent the mean values of triplicates. White bars indicate gene expression in the mESCs and gray bars indicate gene expression in the mEpiSCs. (B) WB analysis for Cdh1 and Cdh2 in mESCs and mEpiSCs. (C) Immunofluorescent staining for Cdh1 and Cdh2 in mESCs and mEpiSCs. The scale bars are 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4588589&req=5

f1: Cdh2 is the major cadherin-type expressed by mEpiSCs.(A) qRT-PCR analysis for classical and atypical cadherins in mESCs and mEpiSCs. Bars represent the mean values of triplicates. White bars indicate gene expression in the mESCs and gray bars indicate gene expression in the mEpiSCs. (B) WB analysis for Cdh1 and Cdh2 in mESCs and mEpiSCs. (C) Immunofluorescent staining for Cdh1 and Cdh2 in mESCs and mEpiSCs. The scale bars are 100 μm.
Mentions: We first analyzed the expression of a variety of classical and atypical cadherin genes: Cdh1 (Epithelial-cadherin), Cdh2 (Neuronal-cadherin), Cdh3 (Placental-cadherin), Cdh4 (Retinal-cadherin), Cdh5 (Vascular Endothelial-cadherin), Cdh12 (Neuronal-cadherin II), Cdh13 (T-cadherin, heart-cadherin) and Cdh15 (Myotubular-cadherin) by quantitative RT-PCR (qRT-PCR). In mESCs, Cdh1 was the most highly expressed of the cadherin genes, although Cdh3 was also expressed. In contrast, mEpiSCs predominantly expressed Cdh1 and Cdh2 (Fig. 1A). We next determined Cdh1 and Cdh2 protein expression in mESCs and mEpiSCs by Western blot (WB) analysis (Fig. 1B). Cdh1 was abundant in mESCs, with only low levels of Cdh2. In contrast, Cdh2 was abundant in mEpiSCs, with almost no Cdh1. Immunofluorescence confirmed the expression of Cdh1 and Cdh2 in the mESCs and mEpiSCs, respectively (Fig. 1C). From these results, we conclude that Cdh1 is a mESC status-specific cadherin and that Cdh2 is a mEpiSC status-specific cadherin.

Bottom Line: Western blot analysis and co-immunoprecipitation assays revealed that Cdh2 stabilizes FGFR1 in mEpiSCs.Furthermore, stable transfection of mESCs with Cdh2 cDNA followed by FGF2 supplementation accelerated cell differentiation.Thus, Cdh2 contributes to the establishment and maintenance of FGF signaling-dependent self-renewal in mEpiSCs through stabilization of FGFR1.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell Biology for Regenerative Medicine, Institute of Advanced Clinical Medicine, Kindai University Faculty of Medicine, 377-2 Ohnohigashi, Osaka-sayama, Osaka, Japan 5898511.

ABSTRACT
The cell adhesion molecule Cadherin 2 (Cdh2) plays important roles in somatic cell adhesion, proliferation and migration. Cdh2 is also highly expressed in mouse epiblast stem cells (mEpiSCs), but its function in these cells is unknown. To understand the function of Cdh2 in mEpiSCs, we compared the expression of pluripotency-related genes in mEpiSCs and mouse embryonic stem cells (mESCs) after either Cdh2 knockdown or Cdh2 over-expression. Introduction of specific siRNA against Cdh2 led to attenuation of pluripotency-related genes. Pluripotent gene expression was not recovered by over-expression of Cdh1 following Cdh2 knockdown. Western blot analysis and co-immunoprecipitation assays revealed that Cdh2 stabilizes FGFR1 in mEpiSCs. Furthermore, stable transfection of mESCs with Cdh2 cDNA followed by FGF2 supplementation accelerated cell differentiation. Thus, Cdh2 contributes to the establishment and maintenance of FGF signaling-dependent self-renewal in mEpiSCs through stabilization of FGFR1.

No MeSH data available.


Related in: MedlinePlus