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. 2016 May 24;7(21):31322-35.
doi: 10.18632/oncotarget.8900.

TET1 inhibits gastric cancer growth and metastasis by PTEN demethylation and re-expression

Yao-Fei Pei  1 , Ran Tao  2 , Jian-Fang Li  1 , Li-Ping Su  1 , Bei-Qin Yu  1 , Xiong-Yan Wu  1 , Min Yan  1 , Qin-Long Gu  1 , Zheng-Gang Zhu  1 , Bing-Ya Liu  1
Affiliations

TET1 inhibits gastric cancer growth and metastasis by PTEN demethylation and re-expression

Yao-Fei Pei et al. Oncotarget. .

Abstract

Ten-Eleven Translocation 1 (TET1) is a member of ten eleven translocation enzymes, which convert 5-methylcytosine (5-mC) to 5-hydroxymethylcytosine (5-hmC). TET1 can promote CpG islands demethylation in specific genes and often absent in various cancers. Herein, we found that TET1 expression and 5-hmC content were low in gastric tumors compared to its adjacent non-tumor tissues. Cell proliferation, migration and invasion were enhanced upon TET1 knockdown in gastric cancer cells in vitro. This phenomenon was confirmed by an animal xeongraft model. We also found that TET1 directly binds to the promoter region of PTEN and activates its transcription through demethylation of CpG islands. TET1 knockdown activated AKT and FAK pathways, which were suppressed by PTEN. The activation of AKT and FAK facilitated tumor migration, invasion and accelerated cell growth. In conclusion, we found a novel mechanism that TET1 suppresses tumor cell growth, migration and invasion through demethylation of CpG island in PTEN promoter by increasing 5-hmC content. The re-expressed PTEN subsequently down regulates AKT and FAK activity.

Keywords: 5-hydroxymethylcytosine; 5-methylcytosine; PTEN; TET1; gastric cancer.

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Conflict of interest statement

The authors declared no conflict of interest.

Figures

Figure 1
Figure 1. TET1 is down-regulated in gastric cancer
A. TET1, B. TET2 and C. TET3 mRNA expression in gastric cancer tissues verse adjacent non-tumor tissues. Data are showed as log2 fold changes (tumor/normal). Results were analyzed by the Wilcoxon Signed Ranks test. D. 5-hmC and 5-mC content were detected by dot blot in gastric cancer tissues compared to adjacent normal tissues. E. IHC analysis of TET1 in gastric cancer tissues and adjacent non-tumor tissues (upper panel 20×, lower panel 200×). F. Kaplan–Meier analysis of overall survival in 80 patients with gastric cancer.
Figure 2
Figure 2. TET1 inhibits cell proliferation, migration and invasion in gastric cell lines
A. TET1 knockdown accelerates (left) and over-expression decelerates (right) cell proliferation. Cell proliferation was measured by CCK-8. B. Wound healing assay of TET1 knockdown in NCI-N87 and TET1 over-expression in SGC-7901. C. TET1 knockdown accelerates and over-expression inhibits cell metastasis ability. * p<0.05; ** p<0.01.
Figure 3
Figure 3. TET1 suppresses tumor progress by up regulating PTEN
A & B. TET1 increased mRNA levels of PTEN, p53, hMLH1 and IRX1. C. TET1 knockdown increased p-AKT and p-FAK (left), while PTEN re-expression decreased p-AKT and p-FAK level (right). D. Would healing assay of rescued effect by PTEN re-expression in TET1 KD cells. E. PTEN re-expression in TET1 KD cells inhibited cell migration and invasion. * p<0.05; ** p<0.01.
Figure 4
Figure 4. TET1 activates PTEN transcription mediated by its catalytic domain
A. qPCR analysis of TET1 expression level in KD3 (upper panel) and SGC-7901 (lower panel) cells after over-expression of wild type or mutant TET1. B. Over-expression of mutant TET1 has no effect on PTEN, p53, hMLH1 and IRX1. C. Over-expression of wild type but not mutant TET1 increases 5-hmC and PTEN level, decreases p-AKT and p-FAK in KD3 cell. D. Over-expression of wild type but not mutant TET1 increases PTEN level, decreases p-AKT and p-FAK in SGC-7901 cell. E & F. Over-expression of wild type but not mutant TET1 inhibits cell migration and invasion. * p<0.05; ** p<0.01.
Figure 5
Figure 5. TET1 increases 5-hmC content in the promoter region of PTEN and promotes its demethylation
A & B. TET1 knockdown in NCI-N87 cell increases PTEN promoter methylation. TET1 over-expression in SGC-7901 cell decreases PTEN promoter methylation. C. TET1 knockdown decreases 5-hmC content in PTEN promoter. D. The position of ChIP primers and PCR results of ChIP. * p<0.05; ** p<0.01.
Figure 6
Figure 6. TET1 suppresses gastric cancer cell metastasis and growth in vivo
A. Subcutaneous tumor formation of KD1 and NC cells. B. Tumor growth curves and tumor weight. C. TET1 knockdown accelerated tumor peritoneal spreading. D. Ki-67 IHC analysis of tumors implanted in nude mice. * p<0.05; ** p<0.01.
Figure 7
Figure 7. TET1 increases other canonical tumor suppressor genes
A & B. TET1 knockdown increases promoter methylation of hMLH1 and IRX1 but not p53 in NCI-N87 cell. C. TET1 knockdown decreases 5-hmC content in hMLH1 and IRX1 promoter but not p53. D & E. TET1 over-expression decreases promoter methylation of hMLH1 and IRX1 but not p53 in SGC-7901 cell. F. TET1 over-expression increases 5-hmC content in hMLH1 and IRX1 promoter but not p53.
Figure 8
Figure 8. Schematic presentation of TET1 action in metastasis
See this image and copyright information in PMC

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