Proteomics, as the leading technology in the postgenomic era, plays an important role in screening diagnostic and therapeutic markers for many human malignancies . In this analysis, the "sandwich" sampling method was adopted to ensure the accuracy of the histopathological results of the sampling. Cervical SCC and paired adjacent cervical tissues with the same genetic backgrounds and high comparability were used as the experimental and control groups, respectively. DEPs identified in this way could relatively objectively reflect the process of tumorigenesis and progression of CC. In this study, a total of 7811 proteins were identified through quantitative proteomics. The number of proteins identified in this study was large, and the quality of proteome detection was satisfactory.
In this study, 97 out of 129 DEPs were found to be related to tumorigenesis and the development of human malignancies, including 88 upregulated and 8 downregulated proteins. KEGG pathway analysis showed that 3 DEPs—HRAS, DUSP7, and PLD1— are RAS pathway components. WB and IHC staining analyses consistently confirmed the results of the quantitative proteomic analysis indicating that the HRAS, P-ERK1/2, and PLD1 levels were increased while the DUSP7 level was decreased in CC tissue compared with the paired normal paracancerous tissue. The RAS/RAF/ERK1/2 signaling pathway which involves members of the mitogen-activated protein kinase (MAPK) family, is pivotal in cell signaling networks . The RAS/RAF/MEK/ERK pathway can trigger a series of cascade reactions, namely, protein phosphorylation, amplification of upstream molecular signals and transduction of the signal into the nucleus, thus activating transcription, promoting gene expression, and stimulating infinite cell . RAS has 4 isoforms: HRAS, NRAS, KRAS4A and KRAS4B. HRAS gene overexpression can specifically activate RAF/MEK/ERK and accelerate the G1/S phase transformation of CC cells . MEK/ERK activation is associated with CC cell resistance to cisplatin . However, when a large number of phosphorylated ERKs accumulate in the ucleus, the use of the MEK-specific blocker U0126 cannot reverse the resistance response of ovarian cancer cells to cisplatin . Exploring the regulatory mechanism of ERK1/2 and reversing its phosphorylation are future areas of focus for oncologists.
Based on our data, the decreased expression of DUSP7 and increased expression of PLD1 were significantly associated with a tumor size > 2 cm and parametrial infiltration. In addition, increased expression of p-ERK1/2 and PLD1and decreased expression of DUSP7 in the CC tissue array were adversely related to patients relapse and survival. These results indicate that both DUSP7 and PLD1 have important regulatory roles in the tumorigenic effect of p-ERK1/2 in CC. DUSP7 is a member of the dual specificity phosphatase (DUSP) family. As a negative regulator of MAKP, DUSPs are involved in cell growth, differentiation, proliferation, migration, apoptosis and tumor formation [17, 18]. Many studies have confirmed that DUSPs are related to tumorigenesis and development. DUSP4 is considered a candidate tumor suppressor gene, and its deletion is related to the occurrence of breast cancer, rectal cancer, thyroid cancer and other tumors [19–21]. DUSP6 is expressed at low levels in the tumor tissues of many human malignancies, including ovarian cancer and endometrial cancer [22–24]. DUSP1 plays different roles in human tumorigenesis: it acts cancer-promoting factor in lung cancer, and leukemia [25–27], and as a tumor suppressor in head and neck SCC, prostate cancer, and urothelial bladder cancer [28–30].
DUSP7 gene is located on human chromosome 3p21 . DUSP7 has a MAP kinase-binding domain/kinase-acting region that can specifically bind to p-ERK1/2, thus leading to its dephosphorylation. In this way, DUSP7 can promote the meiosis of oocytes , the loss of pluripotency in embryonic stem cells , and the differentiation of T cell . Cooperation with DUSP6 and DUSP9 promotes the development of the middle ear and outer ear in mice . However, the role of DUSP7 in the development of human tumors is still poorly understood and controversial. In 2003, Nissenbaum and his colleagues [35–37] found that DUSP7 was upregulated in peripheral blood mononuclear cells and bone marrow in patients with acute leukemia. However, a more recent study demonstrated that DUSP7 is a tumor suppressor gene. DUSP7 deletion has been identified in a variety of human mesothelioma cells  and tumor tissues . DUSP7 activation can effectively block the cell cycle of (human or mouse) BRCA2-deficient cells and significantly inhibit proliferative activity . Ham et al.  demonstrated that constitutive DUPS6 and DUSP7 expression is inversely related to the expression of inducible DUSPs and the phosphorylation of ERK1/2 in lipopolysaccharide (LPS)-stimulated microglia. DUSP7 downregulation is associated with poor survival in patients with breast cancer . However, the relationship between DUSP7 and tumorigenesis of CC has not been reported in the literature.In this analysis, high expression of DUSP7 significantly inhibited the proliferation, invasion and migration ability and EMT process of SIHA cervical cancer cells. The tumorigenesis ability of SIHA cells in nude mice was also inhibited in this way. In contrast, when DUSP7 expression was decreased, the anchorage-independent growth of SIHA cells was significantly increased. In addition, when DUSP7 mRNA levels were up- or downregulated, the expression of HRAS and p-ERK1/2 in SIHA cells was significantly reduced or increased, respectively. ERK inactivation arrested cells at late G1 phase and not prevented cells from entering S phase and from transitioning from G2 to M phase [43, 44]. Persistent ERK 1/2 phosphorylation is the key for signal transmission from surface receptors to the nucleus, and its continuous activation ultimately promotes cell proliferation and malignant transformation . These data indicated that the biological function of DUSP7 is possibly achieved through dephosphorylation of ERK1/2 and inactivation of the RAS pathway. The expression level of PLD1 is not affected by the upregulation or downregulation of DUSP7, and the effect of PLD1 on the progression of cervical cancer is not dependent on the activity status of DUSP7. The related mechanism remains unclear and warrants additional research.