Effects of DACT2 on epithelial mesenchymal transition of glioma cells and its potential mechanism / 西安交通大学学报(医学版)

Objective: To investigate the effects and potential mechanism of tumor suppressor dishevelled-binding antagonist of beta-catenin 2 (DACT2) on epithelial-mesenchymal transition of glioma cells. Methods: The expressions of DACT2 in glioma cells U87, U251, A172 and SHG44 were detected by RT-PCR after 5-Aza treatment. The methylation status of DACT2 promoter was detected by methylation specific PCR (MSP). Western blot was used to detect the expression of DACT2 protein. U87 cells overexpressing DACT2 were constructed and verified by Western blot. Transwell assay was used to detect cell migratory and invasive ability. The protein levels of E-cadherin, Vimentin, MMP2 and Wnt/β-cadherin pathway proteins, i.e., active-β-cadherin, p-β-cadherin and total β-cadherin, in cells were detected by Western blot. Results: DACT2 expression was observed in all these cells after 5-Aza treatment; untreated U87 and U251 cells did not express DACT2 while A172 and SHG44 cells showed weak expression. The DACT2 promoter was completely methylated in U87 and U251 cells, and partially methylated in A172 and SHG44 cells. The level of DACT2 in U87 and U251 cells was lower than that in A172 and SHG44 cells (P<0.05). After transfection of pcDNA3.1-DACT2, the expression of DACT2 in U87 cells increased significantly (P<0.05), U87 cells overexpressing DACT2 were successfully constructed. Overexpression of DACT2 could significantly inhibit epithelial-mesenchymal transition, invasion and migration of U87 cells and block Wnt/β-catenin pathway activation (P<0.05). Conclusion: The DACT2 promoter in glioma cells is highly methylated, and the exogenous overexpression of DACT2 can promote the epithelial mesenchymal transition, invasion and migration of U87 cells. The underling mechanism may be related to the regulation of Wnt/β-catenin pathway by DACT2.

Effects of DACT1 methylation status on invasion and metastasis of nasopharyngeal carcinoma

BACKGROUND: The purpose of the present study was to investigate the role of the methylation status of the DACT1 gene on the invasion and metastasis of nasopharyngeal carcinoma cells. METHODS: The levels of methylation and expression of the DACT1 gene in nasopharyngeal carcinoma tissues and CNE2 cells were determined by methylation-specific PCR and RT-PCR, respectively. CNE2 cells were treated with 5-aza-2-deoxycytidine, and the variation in the methylation status of the DACT1 gene was detected, as well as the influence of methylation on invasiveness of nasopharyngeal carcinoma cells. RESULTS: The DACT1 gene was hyper-methylated in 44 of 62 cases of nasopharyngeal carcinoma. The DACT1 gene was hyper-methylated in 32 of 38 cases of nasopharyngeal carcinoma with lymph node metastasis, and the DACT1 gene was hyper-methylated in 7 of 24 cases of nasopharyngeal carcinoma without lymph node metastasis. The DACT1 mRNA level was weakly expressed or not expressed in all nasopharyngeal carcinoma tissues with hyper-methylated DACT1 genes; however, the DACT1 mRNA level was highly expressed in nasopharyngeal carcinoma tissues with low expression of the methylated DACT1 gene. The DACT1 gene was hyper-methylated and not expressed in CNE2 cells that did not have 5-aza-2-deoxycytidine treatment. After 5-aza-2-deoxycytidine treatment, the DACT1 gene was demethylated and the expression of DACT1 was restored. Moreover, the invasion ability was inhibited in CNE2 cells treated with 5-aza-2-deoxycytidine. CONCLUSION: The expression of DACT1 was related to the methylation status. High expression of DACT1 may inhibit the invasion and metastasis of nasopharyngeal carcinoma cells.

Mutation analysis on DACT1 gene in children with neural tube defects in northern Chinese Han population / 天津医药

Objective To investigate the correlation between neural tube defects (NTDs) and DACT1 gene, and provide the basic data for disease diagnosis and genetic counseling. Methods Blood samples were obtained from 163 NTDs patients and 480 unrelated healthy individuals. Mutation detection of DACT1 gene and DNA direct sequencing was carried out by PCR amplification. Bioinformatics analysis of these mutated loci was performed. Results Six mutations were found in NTDs patients, including 4 missense mutations (p.R45W, p.D142G, p.N356K and p.V702G). But these mutations were not found in 480 healthy individuals. Three mutated amino acid residues (p.45R, p.142D and p.356N) were highly conservative in evolution, and the mutated carriers were female patients, and suffered from anencephaly. Conclusion DACT1 gene mutation may be a risk factor of NTDs in Han population of northern China.

DACT2 gene promoter area methylation status and mRNA expression in renal cell carcinoma / 重庆医学

Objective To explore the role of the DACT2 gene in the occurrence and development of renal cell carcinoma(RCC).Methods The samples of RCC tissues and corresponding tumor-adjacent tissues after radical operation and normal kidney tissues were collected.The methylation specific PCR (MSP) and real time fluorescence reverse transcriptase-PCR (RT-PCR) methods were adopted to detect the methylation status and mRNA expression of DACT2.The streptavidin-peroxidase (SP) method labeled by immunohistochemistry peroxidase was used to examine the expression of β-catenin protein.Then the relationship between DACT2 gene methylation status and mRNA expression with the clinicopathologic characteristics was analyzed.The relationship between DACT2 gene methylation with mRNA and β-catenin expression was analysed,as well.Results The DACT2 mRNA relative expression level in RCC tissues was 0.427±0.025,which was significantly lower than (0.801±0.047) in tumor-adjacent tissues and (0.872±0.022) in normal tissue,the positive rate of DACT2 gene methylation in RCC tissues was 45.76%,which was significantly higher than 6.78% in tumor-adjacent tissues and 5.08% in normal tissues,the difference was statistically significant (P0.05).The DACT2 gene mRNA expression level in RCC tissues and promoter area methylation occurrence rate had no obvious correlation with the clinical data such as patients age,gender,tumor size,clinical stage and Fuhrman grade (P>0.05).The DACT2 gene mRNA relative level in the methylation group was lower than that in the non-methylation group,the difference was statistically significant (P<0.05).The expression rate of β-catenin protein in cytoplasma in RCC tissues was higher than that in the tumor-adjacent tissues and normal tissues,the difference was statistically significant (P<0.05),moreover,DACT2 gen methylation had a positive correlation with β-catenin protein expression (r=0.324,P=0.012).Conclusion The decrease of DACT2 gene promoter area methylation and mRNA relative expression level may participate in the RCC occurrence,but has no relationship with RCC clinical progression.Methylation occurred in DACT2 gene promoter area may be one of reasons causing mRNA relative expression decrase.DACT2 gene methylation occurrence in RCC tissue might be related to the high expression of β-catenin.

Dact genes involved in wnt and TGF-β signaling pathway and their relationship with tumorigenesis / 国际外科学杂志

Dact proteins belong to a structure-related protein family. Recent studies have demonstrated that Dact proteins play an important role in tumorigenesis via modulation of wnt and TGF-p signaling. Delin-eation of the physiological function of Dact proteins would enhance our understanding of look for new strategy targets for cancer and suggests a potential strategy for therapeutic control of wnt and TGF-β signaling in canc-er.