The presence of high-risk human papillomavirus (HR-HPV) types, such as 16 and 18, was a well-established cause in the development of cervical carcinomas . Of course, only a small percentage of HPV-infected patients suffered from cervical cancer, and other cofactors may served to the development of cervical cancers. Therefore, looking for factors other than HPV infection has become a hot spot in the research of cervical cancer. Studies have found that UPP was closely related to the occurrence and development of tumors in some female-related diseases. Studies related to cervical cancer originated from HPV virus, and p53 was degraded through the ubiquitin-proteasome pathway [11, 12], leading to cells cycle changes and loss of apoptotic function. As an upstream component of the UPS, RNF6, a E3 ligase, has attracted more and more attention in the tumor field. Growing evidence has emerged indicating that overexpression of RNF6 was observed in cervical cancer and could develop distant metastasis to a great extent, which ultimately lead to a poor prognosis in CC patients.
However, the role in cancer pathogenesis played by RNF6 remain mysterious. The function and clinical role of RNF6 expression in cervical cancer is has not yet been fully elucidated. In our research, RNF6 acted as an oncogene in cervical cancer. To characterize the expression pattern of RNF6, we performed immunohistochemistry in 20 cervical cancer tissues and western blot in HeLa cells. Consistent with the data from GEPIA, RNF6 protein expression levels is substantially increased compared with normal tissues, which indicated that RNF6 expression participated in the tumorigenesis of cervical cancer. Moreover, the data from GEPIA showed that the clinical prognosis of patients with RNF6-positive staining is significantly poor. According to multivariate analysis, RNF6 overexpression was considered to act as an independent prognostic indicator. In contrast, RNF6 was first found mutated on chromosome 13q12 and performed as a tumor suppressor in human esophageal squamous cell carcinoma . Along with these data, our study confirmed that RNF6 may exert variable or even opposing roles in different tumor contexts and could be expected to be a novel candidate for cancer immunotherapy.
Proliferation of cancer cells is one of signs of malignant tumor and dedicates a poor prognosis of cancer patients. In our study, we designed two effective small interfering RNA (siRNA) sequences to knock down endogenous RNF6 expression and RNF6 overexpression plasmid to enforce the RNF6 expression in cervical cancer HeLa cells to detect the effects of RNF6 in order to obtain a greater understanding of the potential association between the cell-biological characters and the expression levels of RNF6. In vitro studies suggested that RNF6 is capable of inhibiting cell apoptosis to promote cell proliferation, consistent with the results obtained in tumor tissues. The above results indicated that RNF6 exerted its oncogenic role through promoting proliferation and suppressing apoptosis of CC cells.
MAPK/ERK pathway functions a lot in regulating various physiological and pathophysiological processes in the initiation and progression of different types of cancers[13, 14], and ERK is a key member, which can transmit various extracellular stimulation signals into the cell. In addition, most studies currently believe that abnormal activation of ERK signaling pathway can inhibit cell apoptosis in multiple aspects and promote tumor cell growth. Recent studies have demonstrated that RNF135 and RNF138 amplification activated ERK signaling pathway in glioblastoma, which promoted cellular proliferation, migration and invasion [16, 17]. Ruan et al. found that ERK was over expression in cervical cancer. And during the development from CINI to CIN III, the MAPK/ERK pathway was significantly activated . This conclusion was consistent with the research by Li Gang et al., which suggested that the MAPK/ERK signaling pathway played an important role in the evolution of cervical cancer and could promote the cancerous tendency of cervical cells . Therefore, we explored whether the high expression of RFN6 could promote cell proliferation together with its possible molecular mechanisms.
The activation of the ERK signaling pathway requires the activation of ERK by phosphorylation, which can mediate the transmission of signals from the cytoplasm to the nucleus, by up- or down-regulating nuclear transcription factors, such as c-myc, c-Fos, c-Jun, cyclin-D1 and Bcl-2 . c-Fos and c-Jun, as oncogenes encoding nuclear proteins, are highly expressed in cervical cancer.And it has been demonstrated that ERK may affect Hela cell proliferation by regulating the expression of c-Fos and c-Jun proteins . In present study, we performed western blot analysis to detect the expression of p-ERK1/2 and total ERK1/2 when enforced RNF6 expression. While the former was notably elevated, the latter were not altered. Given these evidences, we speculated that we could knocked down RNF6 accompanied by inhibition of MAPK/ERK signaling pathway as well as its downstream genes to treat cervical cancer. The intrinsic mechanisms involved remain unknown and still require further study.
However, our research had some limitations that future research should address. Due to inadequate funding, in vitro and animal experiments have not yet been implemented to determine the specific molecular mechanisms of RNF6-ERK signaling. When we get enough support, we would continue to perfect this deficiency in future experiments.
In summary, we confirmed that RNF6 expression was significantly upregulated in cervical cancer samples and HeLa cell lines than in normal tissues, which also facilitated a poor prognosis for cervical cancer patients. At the same time, RNF6 knockdown might inhibit cell proliferation and promote cell apoptosis by suppression of MAPK/ERK signaling pathway in cervical cancer. Our research has provided a direction in cervical cancer and might important information for better understanding of the mechanics of RNF6-related cervical carcinogenesis. Further experimental studies for RNF6 could develop better therapeutic strategies for cervical cancer.