N6-methyladenosine modification of CENPK mRNA by ZC3H13 promotes cervical cancer stemness and chemoresistance

Abstract

Background Stemness and chemoresistance contribute to cervical cancer recurrence and metastasis. In the current study, we determined the relevant players and role of N6-methyladenine (m6A) RNA methylation in cervical cancer progression. Methods The roles of m6A RNA methylation and centromere protein K (CENPK) in cervical cancer were analyzed using bioinformatics analysis. Methylated RNA immunoprecipitation was adopted to detect m6A modification of CENPK mRNA. Human cervical cancer clinical samples, cell lines, and xenografts were used for analyzing gene expression and function. Immunofluorescence staining and the tumorsphere formation, clonogenic, MTT, and EdU assays were performed to determine cell stemness, chemoresistance, migration, invasion, and proliferation in HeLa and SiHa cells, respectively. Western blot analysis, co-immunoprecipitation, chromatin immunoprecipitation, and luciferase reporter, cycloheximide chase, and cell fractionation assays were performed to elucidate the underlying mechanism. Results Bioinformatics analysis of public cancer datasets revealed firm links between m6A modification patterns and cervical cancer prognosis, especially through ZC3H13-mediated m6A modification of CENPK mRNA. CENPK expression was elevated in cervical cancer, associated with cancer recurrence, and independently predicts poor patient prognosis [hazard ratio = 1.413, 95% confidence interval = 1.078 − 1.853, P = 0.012]. Silencing of CENPK prolonged the overall survival time of cervical cancer-bearing mice and improved the response of cervical cancer tumors to chemotherapy in vivo (P < 0.001). We also showed that CENPK was directly bound to SOX6 and disrupted the interactions of CENPK with β-catenin, which promoted β-catenin expression and nuclear translocation, facilitated p53 ubiquitination, and led to activation of Wnt/β-catenin signaling, but suppression of the p53 pathway. This dysregulation ultimately enhanced the tumorigenic pathways required for cell stemness, DNA damage repair pathways necessary for cisplatin/carboplatin resistance, epithelial-mesenchymal transition involved in metastasis, and DNA replication that drove tumor cell proliferation. Conclusions CENPK was shown to have an oncogenic role in cervical cancer and can thus serve as a prognostic indicator and novel target for cervical cancer treatment.

Comprehensive characterization of tumor microenvironment and m6A RNA methylation regulators and its effects on PD-L1 and immune infiltrates in cervical cancer

“…ZC3H13 expression was shown to be drastically reduced in endometrial carcinoma tissues in a prior study, and it was found to inhibit endometrial carcinoma cell lines from increasing proliferation and invasion (34). ZC3H13 enhanced stemness and chemoresistance via modulation of CENPK mRNA in cervical cancer cells (35). However, the relevance of METTL16 and ZC3H13 in cervical cancer is still not fully known.…”

RNA m6Amethylation in cancer

“…Lin et al. [ 231 ] found that ZC3H13‐mediated m 6 A modification of CENPK mRNA promoted cervical cancer stemness and chemoresistance (Fig. 6B ).…”

“…6A). Lin et al [231] found that ZC3H13-mediated m 6 A modification of CENPK mRNA promoted cervical cancer [204]. (B) METTL14-mediated m 6 A modification on THBS1 mRNA promotes YTHDF2-mediated THBS1 decay in PRAD cells [205].…”

“…(A) METTL3 methylates TFAP2C, which enhances the stability of TFAP2C to increase chemoresistance in seminoma[230]. (B) ZC3H13 targets m6 A on CENPK to activates Wnt/p53 signaling and therefore enhances chemoresistance in cervical cancer[231]. (C) m 6 A-modified ESRRG mRNA upregulates protein expression of both ABCB1 and CPT1B to enhance chemoresistance in BC, LC and HCC cells[232].…”

Recent developments in application of single-cell RNA sequencing in the tumour immune microenvironment and cancer therapy

“…Traditional bulk genomic and transcriptomic analyses average signals from a group of different cells, which obscures the identification of specific cell types and states. In situ hybridization and immunohistochemistry have been used to explore the genomic, transcriptomic, and proteomic characteristics of individual cells, but their outputs are relatively low [ 15 , 16 ]. Flow cytometry and cytometry by time-of-flight (CyTOF) are capable of analysing thousands or millions of single-cell proteomic profiles [ 17 ]; however, these methods require prior selection of antibodies of interest.…”