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DDX3 promotes tumor invasion in colorectal cancer via the CK1ε/Dvl2 axis.

He TY, Wu DW, Lin PL, Wang L, Huang CC, Chou MC, Lee H - Sci Rep (2016)

Bottom Line: The TCF promoter activity and invasion capability were concomitantly increased and decreased by DDX3 manipulation in these cells.The expression of pDvl2 occurred more frequently in high-nuclear than in low-nuclear β-catenin tumors.A prognostic significance of DDX3, pDvl2, and nuclear β-catenin on overall survival and relapse free survival was observed in this study population.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan.

ABSTRACT
DDX3, a subunit of CK1ε, phosphorylates Dvl2 to promote β-catenin activation. Overexpression of the Dvl2 protein results in potent activation of β-catenin/TCF signaling in colorectal cancer. Therefore, we hypothesized that DDX3 might promote tumor invasion via the CK1ε/Dvl2 axis due to β-catenin/TCF activation. Western blotting showed that β-catenin expression was decreased by DDX3 knockdown and increased by DDX3 overexpression in colorectal cancer cells. The TCF promoter activity and invasion capability were concomitantly increased and decreased by DDX3 manipulation in these cells. The invasion capability in colon cancer cells and xenograft lung tumor nodules induced by a DDX3-overexpressing T84 stable clone in tail-vein injection model were nearly suppressed by inhibitors of CK1ε (PF4800567) and β-catenin/TCF signaling (XAV939). Among colorectal cancer patients, DDX3 expression was positively correlated with the expression of pDvl2 and nuclear β-catenin in tumor tissues. The expression of pDvl2 occurred more frequently in high-nuclear than in low-nuclear β-catenin tumors. A prognostic significance of DDX3, pDvl2, and nuclear β-catenin on overall survival and relapse free survival was observed in this study population. We therefore suggest CK1ε or β-catenin/TCF signaling as potential targets for improving tumor regression and outcomes in colorectal cancer, particularly tumors with high-DDX3/high-nuclear β-catenin or high-DDX3/high-pDvl2/high-nuclear β-catenin expression.

No MeSH data available.


Related in: MedlinePlus

DDX3-mediated β-catenin/TCF activation is responsible for invasiveness in colon cancer cells and for lung tumor nodules formation in nude mice.(a) T84 cells were transfected with the indicated combination of TCF promoter plasmid and DDX3-overexpression plasmid. After 24 h, these cells were treated with a CKlε inhibitor (PF4800567) or a β-catenin inhibitor (XAV939) for an additional 5 h, and then the invasion capability for indicated cells was determined by Boyden chamber assays. The TCF promoter activity was evaluated by a luciferase reporter assay. The invasion ability and the TCF promoter activity of these cells with different treatments were shown as the fold changes compared with their NC and VC. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector (VC) or non-specific shRNA controls (NC). #P < 0.05 compared with DDX3-overexpressing T84 cells. (b) Examples of lungs of mice with visual lung tumor nodules at 6 weeks after tail vein inoculation of indicated cells. Representative hematoxylin and eosin staining demonstrates the lung tumor nodules from each group of mice. The number of lung tumor nodules in each group of mice is shown. β-catenin expressions in lung tumor nodules were evaluated by immunohistochemistry using a specific antibody. Data are presented as means ± SD. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector control. #P < 0.05 compared with DDX3-overexpressing T84 cells. The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary information Figure S2.
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f3: DDX3-mediated β-catenin/TCF activation is responsible for invasiveness in colon cancer cells and for lung tumor nodules formation in nude mice.(a) T84 cells were transfected with the indicated combination of TCF promoter plasmid and DDX3-overexpression plasmid. After 24 h, these cells were treated with a CKlε inhibitor (PF4800567) or a β-catenin inhibitor (XAV939) for an additional 5 h, and then the invasion capability for indicated cells was determined by Boyden chamber assays. The TCF promoter activity was evaluated by a luciferase reporter assay. The invasion ability and the TCF promoter activity of these cells with different treatments were shown as the fold changes compared with their NC and VC. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector (VC) or non-specific shRNA controls (NC). #P < 0.05 compared with DDX3-overexpressing T84 cells. (b) Examples of lungs of mice with visual lung tumor nodules at 6 weeks after tail vein inoculation of indicated cells. Representative hematoxylin and eosin staining demonstrates the lung tumor nodules from each group of mice. The number of lung tumor nodules in each group of mice is shown. β-catenin expressions in lung tumor nodules were evaluated by immunohistochemistry using a specific antibody. Data are presented as means ± SD. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector control. #P < 0.05 compared with DDX3-overexpressing T84 cells. The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary information Figure S2.

Mentions: We next examined whether an inhibitor of CK1ε or of β-catenin/TCF signaling could inhibit the cell invasion capability induced by DDX3 overexpression in T84 cells. Western blotting showed that pDvl2 expression was markedly decreased by treatment of the DDX3-overexpressing T84 cells with the CK1ε inhibitor PF4800567. β-catenin expression was markedly decreased by treatment with PF4800567 or with a β-catenin/TCF signaling inhibitor (XAV939) (Fig. 3a upper panel). The TCF promoter activity and invasion capability were reduced by both inhibitors in DDX3-overexpressing T84 cells, but the inhibitory effect on both activities was slightly greater with XAV939 than with PF4800567 (Fig. 3a lower panel). The 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay showed that cell viability was essentially unchanged by DDX3 manipulation and drug treatments at 16 h (Supplementary Figure 3). The tail-vein injection animal model showed that PF4800567 and XAV939 treatment significantly reduced the number of lung tumor nodules in nude mice injected with the DDX3-overexpressing T84 stable clone (Fig. 3b). Consistent with the cell model, the inhibitory effects on lung tumor nodules formations were greater with XAV939 than with PF4800567 (Fig. 3b). These results obtained from the animal model support the mechanistic action of the cell model and suggest that DDX3-mediated β-catenin/TCF activation occurs partially through the CK1ε/Dvl2 axis.


DDX3 promotes tumor invasion in colorectal cancer via the CK1ε/Dvl2 axis.

He TY, Wu DW, Lin PL, Wang L, Huang CC, Chou MC, Lee H - Sci Rep (2016)

DDX3-mediated β-catenin/TCF activation is responsible for invasiveness in colon cancer cells and for lung tumor nodules formation in nude mice.(a) T84 cells were transfected with the indicated combination of TCF promoter plasmid and DDX3-overexpression plasmid. After 24 h, these cells were treated with a CKlε inhibitor (PF4800567) or a β-catenin inhibitor (XAV939) for an additional 5 h, and then the invasion capability for indicated cells was determined by Boyden chamber assays. The TCF promoter activity was evaluated by a luciferase reporter assay. The invasion ability and the TCF promoter activity of these cells with different treatments were shown as the fold changes compared with their NC and VC. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector (VC) or non-specific shRNA controls (NC). #P < 0.05 compared with DDX3-overexpressing T84 cells. (b) Examples of lungs of mice with visual lung tumor nodules at 6 weeks after tail vein inoculation of indicated cells. Representative hematoxylin and eosin staining demonstrates the lung tumor nodules from each group of mice. The number of lung tumor nodules in each group of mice is shown. β-catenin expressions in lung tumor nodules were evaluated by immunohistochemistry using a specific antibody. Data are presented as means ± SD. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector control. #P < 0.05 compared with DDX3-overexpressing T84 cells. The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary information Figure S2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4759588&req=5

f3: DDX3-mediated β-catenin/TCF activation is responsible for invasiveness in colon cancer cells and for lung tumor nodules formation in nude mice.(a) T84 cells were transfected with the indicated combination of TCF promoter plasmid and DDX3-overexpression plasmid. After 24 h, these cells were treated with a CKlε inhibitor (PF4800567) or a β-catenin inhibitor (XAV939) for an additional 5 h, and then the invasion capability for indicated cells was determined by Boyden chamber assays. The TCF promoter activity was evaluated by a luciferase reporter assay. The invasion ability and the TCF promoter activity of these cells with different treatments were shown as the fold changes compared with their NC and VC. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector (VC) or non-specific shRNA controls (NC). #P < 0.05 compared with DDX3-overexpressing T84 cells. (b) Examples of lungs of mice with visual lung tumor nodules at 6 weeks after tail vein inoculation of indicated cells. Representative hematoxylin and eosin staining demonstrates the lung tumor nodules from each group of mice. The number of lung tumor nodules in each group of mice is shown. β-catenin expressions in lung tumor nodules were evaluated by immunohistochemistry using a specific antibody. Data are presented as means ± SD. The P value was statistically determined by the Student’s t-test. *P < 0.05 compared with vector control. #P < 0.05 compared with DDX3-overexpressing T84 cells. The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary information Figure S2.
Mentions: We next examined whether an inhibitor of CK1ε or of β-catenin/TCF signaling could inhibit the cell invasion capability induced by DDX3 overexpression in T84 cells. Western blotting showed that pDvl2 expression was markedly decreased by treatment of the DDX3-overexpressing T84 cells with the CK1ε inhibitor PF4800567. β-catenin expression was markedly decreased by treatment with PF4800567 or with a β-catenin/TCF signaling inhibitor (XAV939) (Fig. 3a upper panel). The TCF promoter activity and invasion capability were reduced by both inhibitors in DDX3-overexpressing T84 cells, but the inhibitory effect on both activities was slightly greater with XAV939 than with PF4800567 (Fig. 3a lower panel). The 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay showed that cell viability was essentially unchanged by DDX3 manipulation and drug treatments at 16 h (Supplementary Figure 3). The tail-vein injection animal model showed that PF4800567 and XAV939 treatment significantly reduced the number of lung tumor nodules in nude mice injected with the DDX3-overexpressing T84 stable clone (Fig. 3b). Consistent with the cell model, the inhibitory effects on lung tumor nodules formations were greater with XAV939 than with PF4800567 (Fig. 3b). These results obtained from the animal model support the mechanistic action of the cell model and suggest that DDX3-mediated β-catenin/TCF activation occurs partially through the CK1ε/Dvl2 axis.

Bottom Line: The TCF promoter activity and invasion capability were concomitantly increased and decreased by DDX3 manipulation in these cells.The expression of pDvl2 occurred more frequently in high-nuclear than in low-nuclear β-catenin tumors.A prognostic significance of DDX3, pDvl2, and nuclear β-catenin on overall survival and relapse free survival was observed in this study population.

View Article: PubMed Central - PubMed

Affiliation: Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan.

ABSTRACT
DDX3, a subunit of CK1ε, phosphorylates Dvl2 to promote β-catenin activation. Overexpression of the Dvl2 protein results in potent activation of β-catenin/TCF signaling in colorectal cancer. Therefore, we hypothesized that DDX3 might promote tumor invasion via the CK1ε/Dvl2 axis due to β-catenin/TCF activation. Western blotting showed that β-catenin expression was decreased by DDX3 knockdown and increased by DDX3 overexpression in colorectal cancer cells. The TCF promoter activity and invasion capability were concomitantly increased and decreased by DDX3 manipulation in these cells. The invasion capability in colon cancer cells and xenograft lung tumor nodules induced by a DDX3-overexpressing T84 stable clone in tail-vein injection model were nearly suppressed by inhibitors of CK1ε (PF4800567) and β-catenin/TCF signaling (XAV939). Among colorectal cancer patients, DDX3 expression was positively correlated with the expression of pDvl2 and nuclear β-catenin in tumor tissues. The expression of pDvl2 occurred more frequently in high-nuclear than in low-nuclear β-catenin tumors. A prognostic significance of DDX3, pDvl2, and nuclear β-catenin on overall survival and relapse free survival was observed in this study population. We therefore suggest CK1ε or β-catenin/TCF signaling as potential targets for improving tumor regression and outcomes in colorectal cancer, particularly tumors with high-DDX3/high-nuclear β-catenin or high-DDX3/high-pDvl2/high-nuclear β-catenin expression.

No MeSH data available.


Related in: MedlinePlus