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SOX4 inhibits GBM cell growth and induces G0/G1 cell cycle arrest through Akt-p53 axis.

Zhang J, Jiang H, Shao J, Mao R, Liu J, Ma Y, Fang X, Zhao N, Zheng S, Lin B - BMC Neurol (2014)

Bottom Line: High SOX4 expression was significantly associated with good prognosis of primary GBMs. SOX4 inhibited the growth of GBM cell line LN229, A172G and U87MG, partly via the activation of p53-p21 signaling and down-regulation of phosphorylated AKT1.Gene expression profiling and subsequent gene ontology analysis showed that SOX4 influenced several key pathways including the Wnt/ beta-catenin and TGF-beta signaling pathways.Our study found that SOX4 acts as a tumor suppressor in GBM cells by induce cell cycle arrest and inhibiting cell growth.

View Article: PubMed Central - PubMed

Affiliation: Cancer Institute (Key Laboratory of Cancer Prevention and Intervention, China National Ministry of Education), Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, Zhejiang Province, P R China. jingzh1985@gmail.com.

ABSTRACT

Background: SOX4 is a transcription factor required for tissue development and differentiation in vertebrates. Overexpression of SOX4 has been reported in many cancers including glioblastoma multiforme (GBM), however, the underlying mechanism of actions has not been studied. In this study, we investigated the role of SOX4 in GBM.

Methods: Kaplan-Meier analysis was performed to assess the association between SOX4 expression levels and survival times in primary GBM samples. Cre/lox P system was used to generate gain or loss of SOX4 in GBM cells, and microarray analysis uncovered the regulation network of SOX4 in GBM cells.

Results: High SOX4 expression was significantly associated with good prognosis of primary GBMs. SOX4 inhibited the growth of GBM cell line LN229, A172G and U87MG, partly via the activation of p53-p21 signaling and down-regulation of phosphorylated AKT1. Gene expression profiling and subsequent gene ontology analysis showed that SOX4 influenced several key pathways including the Wnt/ beta-catenin and TGF-beta signaling pathways.

Conclusions: Our study found that SOX4 acts as a tumor suppressor in GBM cells by induce cell cycle arrest and inhibiting cell growth.

No MeSH data available.


Related in: MedlinePlus

Overexpression of SOX4 reduces proliferation and colony formation of GBM cells. (A) Validation of SOX4 expression in stable SOX4 overexpression cells (LN229_pSFH/A172_pSFH/U87_pSFH) and control cells (LN229_con/A172_con/U87_con) by Real-time PCR and Western blot with anti-HA antibody. (B) The vitality of GBM cells stably express Flag-SOX4-HA or GFP was determined by CCK8 assay. Values at the indicated time points were provided as the mean cell number with an SD of eight wells. *, P < 0.05 (C) Colonies formed by LN229_pSFH/A172_pSFH or LN229_con/A172_con were shown 2 weeks after plating in 6 well plate. Right panel showed the quantification of the colony formation. Values are the means ± SD of triplicate experiments. *, P < 0.05 (D) Colonies formed by U87_pSFH or U87_con were shown 3 weeks after plating in Soft agar.
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Fig3: Overexpression of SOX4 reduces proliferation and colony formation of GBM cells. (A) Validation of SOX4 expression in stable SOX4 overexpression cells (LN229_pSFH/A172_pSFH/U87_pSFH) and control cells (LN229_con/A172_con/U87_con) by Real-time PCR and Western blot with anti-HA antibody. (B) The vitality of GBM cells stably express Flag-SOX4-HA or GFP was determined by CCK8 assay. Values at the indicated time points were provided as the mean cell number with an SD of eight wells. *, P < 0.05 (C) Colonies formed by LN229_pSFH/A172_pSFH or LN229_con/A172_con were shown 2 weeks after plating in 6 well plate. Right panel showed the quantification of the colony formation. Values are the means ± SD of triplicate experiments. *, P < 0.05 (D) Colonies formed by U87_pSFH or U87_con were shown 3 weeks after plating in Soft agar.

Mentions: Encouraged by our data from the transient analysis, we decided to stably over express SOX4 in GBM cells. In order to easily remove the SOX4 in the overexpression construct as a way to study functional reversal, we decided to use a Cre/lox P recombination system to engineer SOX4 overexpression. GBM cell lines LN229, A172, U87MG were selected to over-express Flag-SOX4-HA by retroviral infection. After selected with 800ng/ml puromycin for three weeks, cells were harvested and their expression of exogenous Flag-SOX4-HA were validated by real-time PCR and western blot (Figure 3A).Figure 3


SOX4 inhibits GBM cell growth and induces G0/G1 cell cycle arrest through Akt-p53 axis.

Zhang J, Jiang H, Shao J, Mao R, Liu J, Ma Y, Fang X, Zhao N, Zheng S, Lin B - BMC Neurol (2014)

Overexpression of SOX4 reduces proliferation and colony formation of GBM cells. (A) Validation of SOX4 expression in stable SOX4 overexpression cells (LN229_pSFH/A172_pSFH/U87_pSFH) and control cells (LN229_con/A172_con/U87_con) by Real-time PCR and Western blot with anti-HA antibody. (B) The vitality of GBM cells stably express Flag-SOX4-HA or GFP was determined by CCK8 assay. Values at the indicated time points were provided as the mean cell number with an SD of eight wells. *, P < 0.05 (C) Colonies formed by LN229_pSFH/A172_pSFH or LN229_con/A172_con were shown 2 weeks after plating in 6 well plate. Right panel showed the quantification of the colony formation. Values are the means ± SD of triplicate experiments. *, P < 0.05 (D) Colonies formed by U87_pSFH or U87_con were shown 3 weeks after plating in Soft agar.
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Related In: Results  -  Collection

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Fig3: Overexpression of SOX4 reduces proliferation and colony formation of GBM cells. (A) Validation of SOX4 expression in stable SOX4 overexpression cells (LN229_pSFH/A172_pSFH/U87_pSFH) and control cells (LN229_con/A172_con/U87_con) by Real-time PCR and Western blot with anti-HA antibody. (B) The vitality of GBM cells stably express Flag-SOX4-HA or GFP was determined by CCK8 assay. Values at the indicated time points were provided as the mean cell number with an SD of eight wells. *, P < 0.05 (C) Colonies formed by LN229_pSFH/A172_pSFH or LN229_con/A172_con were shown 2 weeks after plating in 6 well plate. Right panel showed the quantification of the colony formation. Values are the means ± SD of triplicate experiments. *, P < 0.05 (D) Colonies formed by U87_pSFH or U87_con were shown 3 weeks after plating in Soft agar.
Mentions: Encouraged by our data from the transient analysis, we decided to stably over express SOX4 in GBM cells. In order to easily remove the SOX4 in the overexpression construct as a way to study functional reversal, we decided to use a Cre/lox P recombination system to engineer SOX4 overexpression. GBM cell lines LN229, A172, U87MG were selected to over-express Flag-SOX4-HA by retroviral infection. After selected with 800ng/ml puromycin for three weeks, cells were harvested and their expression of exogenous Flag-SOX4-HA were validated by real-time PCR and western blot (Figure 3A).Figure 3

Bottom Line: High SOX4 expression was significantly associated with good prognosis of primary GBMs. SOX4 inhibited the growth of GBM cell line LN229, A172G and U87MG, partly via the activation of p53-p21 signaling and down-regulation of phosphorylated AKT1.Gene expression profiling and subsequent gene ontology analysis showed that SOX4 influenced several key pathways including the Wnt/ beta-catenin and TGF-beta signaling pathways.Our study found that SOX4 acts as a tumor suppressor in GBM cells by induce cell cycle arrest and inhibiting cell growth.

View Article: PubMed Central - PubMed

Affiliation: Cancer Institute (Key Laboratory of Cancer Prevention and Intervention, China National Ministry of Education), Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, Zhejiang Province, P R China. jingzh1985@gmail.com.

ABSTRACT

Background: SOX4 is a transcription factor required for tissue development and differentiation in vertebrates. Overexpression of SOX4 has been reported in many cancers including glioblastoma multiforme (GBM), however, the underlying mechanism of actions has not been studied. In this study, we investigated the role of SOX4 in GBM.

Methods: Kaplan-Meier analysis was performed to assess the association between SOX4 expression levels and survival times in primary GBM samples. Cre/lox P system was used to generate gain or loss of SOX4 in GBM cells, and microarray analysis uncovered the regulation network of SOX4 in GBM cells.

Results: High SOX4 expression was significantly associated with good prognosis of primary GBMs. SOX4 inhibited the growth of GBM cell line LN229, A172G and U87MG, partly via the activation of p53-p21 signaling and down-regulation of phosphorylated AKT1. Gene expression profiling and subsequent gene ontology analysis showed that SOX4 influenced several key pathways including the Wnt/ beta-catenin and TGF-beta signaling pathways.

Conclusions: Our study found that SOX4 acts as a tumor suppressor in GBM cells by induce cell cycle arrest and inhibiting cell growth.

No MeSH data available.


Related in: MedlinePlus