Limits...
Porf-2 Inhibits Neural Stem Cell Proliferation Through Wnt/β-Catenin Pathway by Its GAP Domain.

Huang GH, Yang XT, Chen K, Xing J, Guo L, Zhu L, Li HJ, Li XC, Zhang SY, Feng DF - Front Cell Neurosci (2016)

Bottom Line: Previous studies showed porf-2 functions as a modulator in central nerve system development.We here show that porf-2, a conserved family of RhoGAPs, is highly and specifically expressed in NSCs.By using neurosphere formation and Edu assay we confirmed the GAP domain is necessary for its function.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine Shanghai, China.

ABSTRACT
Neural stem cell (NSC) proliferation and differentiation play a pivotal role in the development of brain, the plasticity of the brain network, and the repair for brain function in CNS diseases. The mechanisms regulating NSC behavior are not well elucidated. Previous studies showed porf-2 functions as a modulator in central nerve system development. We here show that porf-2, a conserved family of RhoGAPs, is highly and specifically expressed in NSCs. We also demonstrate that porf-2 inhibits the proliferation of NSCs in vivo and in vitro, but has no effect on NSC differentiation. We investigated which domain is required for the role of porf-2 on NSC proliferation. By using neurosphere formation and Edu assay we confirmed the GAP domain is necessary for its function. In addition, we surveyed a few classical pathways on NSC proliferation and found that porf-2 inhibits NSC proliferation by suppressing the β-catenin nuclear translocation. Taken together, we show for the first time that porf-2 inhibits NSC proliferation through Wnt/β-catenin pathway by its GAP domain.

No MeSH data available.


Related in: MedlinePlus

Porf-2 inhibits NSC proliferation through wnt/β-catenin pathway. (A) The expression and quantification of p-PTEN, PTEN, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, NICD, and β-catenin protein in control and porf-2 overexpression group. Data are mean ± SEM (n = 3). β-actin was used as a loading control. (B) The expression and quantification of β-catenin protein in cytoplasm and nucleus in control and porf-2 overexpression group. GAPDH and HistoneH3 were used as loading controls for protein in cytoplasm and nucleus respectively. **p < 0.01. (C) Confocal microscopy image of β-catenin expression in NSCs after lentivirus infection. White arrow indicated the expression of β-catenin in nucleus in control group. Blue arrow indicated the expression of β-catenin in nucleus in porf-2 overexpression group. Scale bar: 50 μm. (D) The expression and quantification of β-catenin protein in cytoplasm and nucleus in different groups. Data are mean ± SEM (n = 3). **p < 0.01.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4814557&req=5

Figure 5: Porf-2 inhibits NSC proliferation through wnt/β-catenin pathway. (A) The expression and quantification of p-PTEN, PTEN, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, NICD, and β-catenin protein in control and porf-2 overexpression group. Data are mean ± SEM (n = 3). β-actin was used as a loading control. (B) The expression and quantification of β-catenin protein in cytoplasm and nucleus in control and porf-2 overexpression group. GAPDH and HistoneH3 were used as loading controls for protein in cytoplasm and nucleus respectively. **p < 0.01. (C) Confocal microscopy image of β-catenin expression in NSCs after lentivirus infection. White arrow indicated the expression of β-catenin in nucleus in control group. Blue arrow indicated the expression of β-catenin in nucleus in porf-2 overexpression group. Scale bar: 50 μm. (D) The expression and quantification of β-catenin protein in cytoplasm and nucleus in different groups. Data are mean ± SEM (n = 3). **p < 0.01.

Mentions: As is well-known, there are a few classical pathways involved in NSC proliferation: PTEN/PI3k-Akt, Akt-mTOR, Notch, and Wnt/β-catenin pathway (Liu and Niswander, 2005; Egeland et al., 2015). To understand the mechanism of how porf-2 inhibited NSC proliferation, we surveyed above pathways. As shown in Figure 6A, p-PTEN/PTEN, p-PI3K/PI3K, p-Akt/Akt protein level remained unchanged after porf-2 overexpression, indicating that porf-2 had no effect on PTEN/PI3K/Akt pathway. Then we found that although p-mTOR protein level had a slight decrease, it showed no significant difference with control (Figure 5A). We also checked the Notch and Wnt/β-catenin pathway and found that there was no difference in NICD and β-catenin protein level between overexpression of porf-2 and control group (Figure 5A). Similar results were obtained when porf-2 was knock down in NSCs (data not shown).


Porf-2 Inhibits Neural Stem Cell Proliferation Through Wnt/β-Catenin Pathway by Its GAP Domain.

Huang GH, Yang XT, Chen K, Xing J, Guo L, Zhu L, Li HJ, Li XC, Zhang SY, Feng DF - Front Cell Neurosci (2016)

Porf-2 inhibits NSC proliferation through wnt/β-catenin pathway. (A) The expression and quantification of p-PTEN, PTEN, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, NICD, and β-catenin protein in control and porf-2 overexpression group. Data are mean ± SEM (n = 3). β-actin was used as a loading control. (B) The expression and quantification of β-catenin protein in cytoplasm and nucleus in control and porf-2 overexpression group. GAPDH and HistoneH3 were used as loading controls for protein in cytoplasm and nucleus respectively. **p < 0.01. (C) Confocal microscopy image of β-catenin expression in NSCs after lentivirus infection. White arrow indicated the expression of β-catenin in nucleus in control group. Blue arrow indicated the expression of β-catenin in nucleus in porf-2 overexpression group. Scale bar: 50 μm. (D) The expression and quantification of β-catenin protein in cytoplasm and nucleus in different groups. Data are mean ± SEM (n = 3). **p < 0.01.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4814557&req=5

Figure 5: Porf-2 inhibits NSC proliferation through wnt/β-catenin pathway. (A) The expression and quantification of p-PTEN, PTEN, p-PI3K, PI3K, p-Akt, Akt, p-mTOR, mTOR, NICD, and β-catenin protein in control and porf-2 overexpression group. Data are mean ± SEM (n = 3). β-actin was used as a loading control. (B) The expression and quantification of β-catenin protein in cytoplasm and nucleus in control and porf-2 overexpression group. GAPDH and HistoneH3 were used as loading controls for protein in cytoplasm and nucleus respectively. **p < 0.01. (C) Confocal microscopy image of β-catenin expression in NSCs after lentivirus infection. White arrow indicated the expression of β-catenin in nucleus in control group. Blue arrow indicated the expression of β-catenin in nucleus in porf-2 overexpression group. Scale bar: 50 μm. (D) The expression and quantification of β-catenin protein in cytoplasm and nucleus in different groups. Data are mean ± SEM (n = 3). **p < 0.01.
Mentions: As is well-known, there are a few classical pathways involved in NSC proliferation: PTEN/PI3k-Akt, Akt-mTOR, Notch, and Wnt/β-catenin pathway (Liu and Niswander, 2005; Egeland et al., 2015). To understand the mechanism of how porf-2 inhibited NSC proliferation, we surveyed above pathways. As shown in Figure 6A, p-PTEN/PTEN, p-PI3K/PI3K, p-Akt/Akt protein level remained unchanged after porf-2 overexpression, indicating that porf-2 had no effect on PTEN/PI3K/Akt pathway. Then we found that although p-mTOR protein level had a slight decrease, it showed no significant difference with control (Figure 5A). We also checked the Notch and Wnt/β-catenin pathway and found that there was no difference in NICD and β-catenin protein level between overexpression of porf-2 and control group (Figure 5A). Similar results were obtained when porf-2 was knock down in NSCs (data not shown).

Bottom Line: Previous studies showed porf-2 functions as a modulator in central nerve system development.We here show that porf-2, a conserved family of RhoGAPs, is highly and specifically expressed in NSCs.By using neurosphere formation and Edu assay we confirmed the GAP domain is necessary for its function.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine Shanghai, China.

ABSTRACT
Neural stem cell (NSC) proliferation and differentiation play a pivotal role in the development of brain, the plasticity of the brain network, and the repair for brain function in CNS diseases. The mechanisms regulating NSC behavior are not well elucidated. Previous studies showed porf-2 functions as a modulator in central nerve system development. We here show that porf-2, a conserved family of RhoGAPs, is highly and specifically expressed in NSCs. We also demonstrate that porf-2 inhibits the proliferation of NSCs in vivo and in vitro, but has no effect on NSC differentiation. We investigated which domain is required for the role of porf-2 on NSC proliferation. By using neurosphere formation and Edu assay we confirmed the GAP domain is necessary for its function. In addition, we surveyed a few classical pathways on NSC proliferation and found that porf-2 inhibits NSC proliferation by suppressing the β-catenin nuclear translocation. Taken together, we show for the first time that porf-2 inhibits NSC proliferation through Wnt/β-catenin pathway by its GAP domain.

No MeSH data available.


Related in: MedlinePlus