br Discussion br This study indicated that ZNF played an
This study indicated that ZNF692 played an important role in pro-moting aggressiveness in CC. Here, we firstly found that the expression of ZNF692 was upregulated in CC tissues compared with its expression in the adjacent normal tissues. Moreover, knockdown or ectopic expres-sion of ZNF692 suppressed or promoted the proliferation and invasion abilities of CC cells, respectively. Furthermore, xenograft tumour models using HeLa Phosphatase Inhibitor Cocktail II showed that knockdown of ZNF692 inhibited tumour growth in vivo. Regarding its molecular mechanism, si-ZNF692 treat-ment of CC cells induced G1 phase arrest, upregulated p27kip1 and
downregulated PThr160-CDK2 expression while overexpression of ZNF692 suppressed p27kip1 and promoted PThr160-CDK2 expression. In-
terestingly, Chromatin immunoprecipitation (ChIP) assay revealed that ZNF692 directly targeted p27kip1 promoter region (815–1022 bp) and
luciferase reporter assay indicated that ectopic or si-ZNF692 signifi-cantly reduced or increased p27kip1 promoter (700-1200 bp) luciferase activity. Additionally, inhibition of p27kip1 expression partially recov-ered the influence of si-ZNF692 on malignant phenotype in Hela cells. Taken together, our results provided evidence that ZNF692 promoted the progression of CC and might be a potential prognostic biomarker as well as the therapeutic target in CC.
The involvement of ZNF692 in cancer has been reported in many studies. For example, ZNF692 was up-regulated in the parous breast via transcription regulation and chromatin organization . Our
previous study demonstrated that down-regulation of ZNF692 could suppress tumour proliferation and invasion of lung adenocarcinoma cells and overexpression of ZNF692 is strongly associated with poorer survival in LUAD . Similar to our previous findings, we found that ZNF692 was overexpressed in CC tissues compared with that in adjacent normal tissues and was negatively correlated with PFS in CC patients. Moreover, overexpression of ZNF692 was also positively correlated with advanced TNM stage and lymph node metastasis based on analysis of CC tissue microarrays.
By GO enrichment analyses, we found that the genes which had the highest correlation values with ZNF692 were enriched in regulation of transcription and the cell cycle. In the past decade, role of cell cycle dys-regulation has been recognized in occurrence and progression of tu-mours [24–26]. There has been mounting evidence to indicate that metabolite profiling has a key role in the study of cancer cell prolifera-tion [27,28] and a better understanding of the metabolic basis of cell cycle disorders is expected to uncover the mechanisms of accelerating tumorigenesis .
Our functional experiments were consistent with the GO analyses, in which knockdown of ZNF692 impaired CC cell proliferation, migration and invasion by inhibiting the G1/S phase transition. Therefore, we analysed several critical G1/S transition-related genes to illuminate a potential mechanism. Previous studies have reported that p27kip1 ar-rests the cell cycle at the G1 phase by inhibiting the activation of the CCNE1-CDK2 complex [30–33]. The distribution and prognostic value
Fig. 4. ZNF692 suppressed the p27kip1 expression by directly binding its promoter region. (A) Genes of high correlation with ZNF692 were enriched in regulation of transcription and cell cycle regulation by GO analysis. (B–D) The p27kip1 mRNA levels were significantly increased or decreased after knockdown or ectopic expression of ZNF692 in CC cell lines, respectively. (E– F) The protein level of p27kip1 was obviously upregulated or downregulated while the expression of PThr160-CDK2 was moderately decreased or increased in si- or oe-ZNF692 of CC cells. The expression of CDK2, CCNE1 was not obviously affected at the protein level after knockdown or ectopic expression of ZNF692 in CC cells. However, the p21cip1, CCND1 and CCNB1 had no significant changes in si- or oe-ZNF692. (G) Each approximately 200 bp length was designed for 7 primers within 1500 bp of the p27kip1 promoter region. Chromatin immunoprecipitation (ChIP) assays using normal IgG or anti-Flag-ZNF692 revealed that ZNF692 directly targeted p27kip1 promoter region (815–1022 bp). (H) Ectopic ZNF692 or si-ZNF692 significantly reduced or increased p27kip1 promoter (700–1200 bp) luciferase activity. Error bars represent the mean ± SD values of three independent experiments. *P b 0.05, **P b 0.01, ***P b 0. 001.