br Classical zinc finger containing proteins ZNFs one
Classical zinc finger-containing proteins (ZNFs), one of the three types of transcription factors, are encoded by 2% of human genes and constitute the largest family of sequence-specific DNA binding proteins [5,6]. Recent research has indicated that aberrant Nocodazole of ZNF pro-teins leads to tumorigenesis in various malignancies [7–9]. Although many studies have investigated the roles of ZNF proteins in various tu-mours, the biological function of ZNF692 has rarely been reported. ZNF692 is located on chromosome 1q44 and can bind to the promoter region of key genes to regulate the tumour progression as a transcrip-tion factor . A previous study indicated that ZNF692 was related to the recurrence of Wilms tumours . Additionally, ZNF692 has differ-ent RNA splicing patterns in various types of hepatocellular carcinoma . Furthermore, our previous research suggests that ZNF692 is upreg-ulated in lung adenocarcinoma (LUAD) and promotes the aggressive-ness of LUAD . Nevertheless, none of these studies have focused on the expression profile and molecular mechanisms of ZNF692 in CC.
To explore the role of ZNF692 in CC, we investigated the expression profiles of ZNF692 in the TCGA dataset, CC tissue microarrays and CC cell lines. Our results suggest that ZNF692 plays an important role in pro-
moting aggressiveness, including proliferation, migration and invasion capacities in CC, by regulating the p27kip1/PThr160-CDK2 signal pathway
to induce the G1/S transition. Thus, ZNF692 might serve as a useful prognostic biomarker and a potential target in cervical carcinoma.
2. Material and methods
2.1. Bioinformatics analysis
A TCGA dataset termed TCGA_CESC_exp_HiS-eqV2-2015-02-24 was downloaded from the UCSC cancer browser (https://genome-cancer. ucsc.edu/) . A total of 292 patients with complete clinical and follow-up information were extracted for further statistical analysis. To-tally 387 genes which had the highest co-expression correlation with ZNF692 (r N 0.4 or r b −0.4) in the TCGA cervical cancer dataset were submitted to DAVID Bioinformatics Resources 6.7 (http://david.abcc. ncifcrf.gov/) [15,16] for Kyoto Encyclopaedia of Genes and Genomes (KEGG) and Gene Ontology (GO) pathway enrichment analysis. And GEPIA (Gene Expression Profiling Interactive Analysis) (http://gepia. cancer-pku.cn/index.html) was used to analyse the expression of ZNF692 with Disease Free Survival (DFS) of CC patients.
2.2. CC tissue microarrays analysis by immunohistochemistry (IHC)
For IHC assays based on tissue microarrays (TMAs), sixty-two pairs of paraffin-embedded human cervical cancer sections were analysed for ZNF692 expression by IHC according to a previous protocol . Paired CC tissue microarrays were obtained from Shanghai Outdo Bio-tech Co., Ltd. (Cat. No. OD-CT-RpUtr 03-004 and OD-CT-RpUtr03-006). And the detailed information of CC cases was shown in Table S1. All tis-sues were examined by two experienced pathologists who confirmed the TNM stage of each patient. The UICC/AJCC TNM staging method (7th edition) was used for clinical staging and the lymph node metasta-sis was confirmed by dissecting lymph nodes during surgery. The sec-tions were incubated with an anti-ZNF692 primary antibody (1:100
dilution; Abcam, ab204595). Tissue microarrays were assessed by two separate investigators blinded to the clinical parameters and then were subjected to statistical analysis. The IHC scores were calculated ac-cording to intensity and percentage of positive cells. The staining inten-sity was evaluated as the basis of 4 grades: (negative staining), 1 (weak staining), 2 (moderate staining), or 3 (strong staining). The prod-uct (percentage of positive cells and respective intensity scores) was used as the final staining scores (a minimum value of and a maximum value of 300).