title: MiR-375 Is Epigenetically Downregulated by HPV-16 E6 Mediated DNMT1 Upregulation and Modulates EMT of Cervical Cancer Cells by Suppressing lncRNA MALAT1.
abstract: Epigenetic modulation is an important mechanism of miRNA dysregulation in cervical cancer. In this study, we firstly studied how this mechanism contributes to miR-375 downregulation in cervical cancer cells. Then, we further studied the association between miR-375 and MALAT1 (metastasis associated lung adenocarcinoma transcript 1) in epithelial mesenchymal transition (EMT) of the cancer cells. HPV-16 positive SiHa and CaSki cells were used as in vitro model. Our data showed that HPV-16 E6 positively modulated DNMT1 expression in both SiHa and CaSki cells. Knockdown of DNMT1 partly restored miR-375 levels in the cells. The following methylation-specific PCR (MSP) assay and qRT-PCR analysis showed that methylation was common in the promoter region of miR-375 in both SiHa and CaSki cells and demethylation partly restored miR-375 levels in the cells. Therefore, we infer that miR-375 is downregulated partly due to promoter hypermethylation mediated by DNMT1 in HPV-16 positive cervical cancer cells. Our bioinformatics analysis showed that MALAT1 has three putative binding sites with miR-375 and the following dual luciferase assay confirmed two of them. QRT-PCR analysis showed that miR-375 overexpression significantly reduced MALAT1 expression, while MALAT1 overexpression reversely suppressed miR-375 levels. Therefore, we infer that there is a reciprocal regulation between miR-375 and MALAT1 in the cells. In SiHa cells, miR-375 overexpression or MALAT1 siRNA partly restored E-cadherin expression, significantly reduced N-cadherin and also reduced invasion capacity of SiHa cells. Therefore, these results suggest that miR-375 and MALAT1 form a functional axis modulating EMT in cervical cancer.
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Shikai
authors.initials: S
authors.lastname: Liu
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Song
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Hairong
authors.initials: H
authors.lastname: Yao
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Dongkui
authors.initials: D
authors.lastname: Xu
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Fangyuan
authors.initials: F
authors.lastname: Gao
authors.affiliation: Department of Obstetrics & Gynaecology, Cangzhou Central Hospital, Hebei, 061001, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Li