Tumor-suppressor gene SOX1 is a methylation-specific expression gene in cervical adenocarcinoma

Abstract

The present study is to analyze the difference of gene methylation in early cervical adenocarcinoma and to find molecular markers for predicting the occurrence and development of cervical adenocarcinoma.A total of 15 cases of primary cervical adenocarcinoma and 10 cases of primary cervical squamous cell carcinoma at stages IB1 or IIA1 were included in the study. Infinium MethylationEPIC BeadChip (850K) was used to screen specifically expressed genes in cervical adenocarcinoma tissues. Bisulfite sequencing polymerase chain reaction (BSP) and quantitative real-time polymerase chain reaction (qRT-PCR) were used to verify the methylation levels in cervical adenocarcinoma, cervical squamous cell carcinoma, and normal cervical tissues.Sex determining region Y-box 1 (SOX1) and cyclin D1 (CCND1) genes participated in multiple signaling pathways, being the central nodes of gene regulatory networks. SOX1 gene, but not CCND1 gene, was a specifically methylated gene in cervical adenocarcinoma according to BSP. According to qRT-PCR, methylation level of SOX1 in cervical adenocarcinoma tissues is significantly different from that in cervical squamous cell carcinoma tissues or normal cervical tissues, and the methylation level of CCND1 in cervical adenocarcinoma tissues or cervical squamous cell carcinoma tissues is significantly different from that in normal cervical tissues.The present study demonstrates that tumor-suppressor gene SOX1 is a methylation-specific expression gene of cervical adenocarcinoma and is expected to become a specific molecular marker for the diagnosis of cervical adenocarcinoma. However, CCND1 gene was not proven to be a specific methylation expression gene in cervical adenocarcinoma in the present study.

Figure 1

Detection of methylation level in cervical adenocarcinoma tissues. (A) Density plot of raw data (753037 probes). (B) Methylation variable position (MVP) analysis between cervical adenocarcinoma tissues and normal cervical tissues.

Figure 2

Methylation levels of promoter regions of SOX1 and CCND1 genes in cervical adenocarcinoma tissues and normal cervical tissues. (A) Total DNA agarose gel electrophoresis. M, marker (100–5000 bp); lanes 1 to 5, normal cervical tissues; lanes 6 to 10, cervical adenocarcinoma tissues; lanes 11 to 15, cervical squamous cell carcinoma tissues. (B) Aggregate plot of methylation status of SOX1 promoter region in cervical adenocarcinoma tissues. (C) Methylation levels of SOX1 promoter region in cervical adenocarcinoma, cervical squamous cell carcinoma and normal cervical tissues. ^∗∗P < .05 compared with normal group. (D) Aggregate plot of methylation status of CCND1 promoter region in cervical adenocarcinoma tissues. (E) Methylation levels of CCND1 promoter region in cervical adenocarcinoma, cervical squamous cell carcinoma, and normal cervical tissues.

Figure 3

Methylation levels of SOX1 and CCND1 in cervical adenocarcinoma, cervical squamous cell carcinoma, and normal cervical tissues. (A) Agarose gel electrophoresis of total RNA. Lanes1 and 2, normal cervical tissues; lanes 3 and 4, cervical adenocarcinoma tissues; lanes 5 and 6, cervical squamous cell carcinoma tissues. (B–D) Amplification and dissociation curves of (B) GAPDH, (C) SOX1, and (D) CCND1. (E, F) Methylation levels of (E) SOX1 and (F) CCND1 in 2 cases of cervical adenocarcinoma tissues, 2 cases of cervical squamous cell carcinoma tissues, and 2 cases of normal cervical tissues.