SEC61G Promotes Cervical Cancer Proliferation by Activating MAPK Signaling Pathway

Abstract

Objective: The abnormal expression of SEC61G plays an important role in the development of various tumors. This study explored the effects of SEC61G on MAPK signaling pathway and proliferation of cervical cancer (CC) cells.

Methods: shRNA was used to inhibit the expression of SEC61G and EdU to observe its effect on the proliferation of CC cell SiHa. The effect of SEC61G on invasion was evaluated by Transwell assay. TCGA database was used to analyze the influence of high or low SEC61G expression level on the overall survival of CC patients. Western blot was used to detect the expressions of SEC61G, p-RAF1, Raf1, p-MEK1/2, MEK1/2, and p-ERK1/2 in cells. SiHa cells overexpressing SEC61G (SiHa-SEC61G) and control group (SiHa-mock) were subcutaneously implanted in nude mice. The tumor growth curve was measured at the specified time points between SiHa-SEC61G and SiHa-mock. The inhibitory effect of gefitinib on SEC61G was further evaluated.

Results: In patients with CC, high SEC61G expression predicted poor prognosis. Silencing SEC61G inhibited proliferation and invasion of CC cells in vitro. Overexpression of SEC61G can promote the proliferation and invasion of CC cells in vitro. Meanwhile, overexpression of SEC61G promoted the proliferation of CC xenografts. Knocking down SEC61G can inhibit MAPK signaling pathway. Gefitinib can inhibit CC proliferation and tumor growth by SEC61G.

Conclusion: SEC61G is highly expressed in CC and has poor prognosis. Inhibition of SEC61G expression can effectively inhibit the growth and proliferation of human CC cells. The mechanism may be related to the inhibition of MAPK signaling pathway.

Figure 1

In CC patients, high SEC61G expression predicts poor prognosis. (a) Kaplan-Meier curve of overall survival in CC patients with high or low SEC61G expression levels in the cohort. The data sources of SEC61G survival analysis were downloaded from The Human Protein Atlas website (https://www.proteinatlas.org/). (b) SEC61G is highly expressed in CC cells. ^∗P < 0.05 and ^∗∗P < 0.01 vs. CerEpiC group.

Figure 2

Silencing of SEC61G inhibits proliferation and invasion of CC cells in vitro. (a) The transfection efficiency of SEC61G was detected by transfecting SiHa (SiHa-shSEC61G) and control group (shNC) with SEC61G knockout shRNA. (b) EdU proliferation assay. (c) Transwell detection of invasive ability of CC cells. After 48 h, crystal violet staining was performed and counted. 200x. (d) Detection of E-cadherin expression. (e) Vimentin expression detection. ^∗P < 0.05, ^∗∗P < 0.01, and ^∗∗∗P < 0.001 vs. shNC group.

Figure 3

Overexpression of SEC61G promotes proliferation and invasion of CC cells in vitro. (a) SiHa cells were transfected with a plasmid overexpressing SEC61G, and the transfection efficiency of SEC61G was detected. (b) EdU proliferation assay. (c) Transwell detection of invasive ability of CC cells. After 48 h, crystal violet staining was performed and counted. (d) Detection of E-cadherin expression. (e) Vimentin expression detection. ^∗P < 0.05 and ^∗∗P < 0.01 vs. mock group.

Figure 4

SEC61G promotes CC growth and progression in vivo. (a) Nude mice were subcutaneously implanted with SiHa cells overexpressing SEC61G (SiHa-SEC61G) and a control group (SiHa-mock). Tumor growth curves were measured at the indicated time points between SiHa-SEC61G and SiHa-mock. ^∗P < 0.05, one-way ANOVA. (b) Each group was injected with 2 × 10⁶ cells; tumors were excised on day 21 postinjection. (c) Tumor weight was measured when the tumor was removed. (d) Representative images of IHC staining showing a positive correlation between Ki-67 and SEC61G overexpression in subcutaneous implants of each group. (e) Relative intensity of Ki-67 staining. ^∗∗P < 0.01 vs. mock group.

Figure 5

Knockdown of SEC61G inhibits MAPK signaling. (a) Western blot detection of key protein expression changes of MAPK signaling pathway. (b) Statistical analysis results of p-RAF1/Raf1. (c) Statistical analysis results of p-MEK1/2-MEK1/2. (d) Statistical analysis results of p-ERK1/2-ERK1/2. ^∗∗P < 0.01 vs. shNC group.

Figure 6

Gefitinib inhibits CC proliferation and tumor growth through SEC61G. (a) After SiHa cells were given gefitinib (10 μM, 24 h) or DMSO, the expression of SEC61G was analyzed by qRT-PCR. (b) EdU proliferation assay. (c) Transwell detection of invasive ability of CC cells. After 48 h, crystal violet staining was performed and counted. (d) Detection of E-cadherin expression e. (e) Vimentin expression detection. (f) Schematic description of the regulatory mechanism of CC proliferation through SEC61G/MAPK signaling and therapeutic targets. ^∗∗P < 0.01 and ^∗∗∗P < 0.001 vs. DMSO group.