The HPV E6-miR-543-CHMP4C Pathway Promotes Tumorigenesis And Progression Of Cervical Cancer

Objective:CHMP4C(Charged multivesicular body protein 4C),as a subunit of endosomal sorting complex required for transport-III(ESCRT-III),participates in the sorting of multiple vesicles(CMV)and plays an important role in the Aurora B kinase-mediated abscission checkpoint,preventing premature cell division and the reduction of DNA damage.Studies have revealed that abnormal expression of CHMP4C could increase the susceptibility of some cancers,such as ovarian cancer,prostate cancer and lung cancer,but its role in cervical cancer has not been clear.This study mainly explored the role of CHMP4C in the tumorigenesis and progression of cervical cancer and the mechanisms it may participate in.Methods:1.Immunohistochemistry was used to detect the expression of CHMP4C in normal cervical and cervical cancer tissues.2.After knocking down the expression of CHMP4C by siRNA in the cervical cancer cell lines SiHa and HeLa,cell proliferation,apoptosis,migration and invasion were examined by MTT assay,plate cloning formation assay,flow cytometry,wound-healing assay and cell invasion assay,respectively.3.Western blotting was used to measure the change of tumor-related regulatory proteins after silencing CHMP4C.4.E6 was down-regulated in HPV16 positive cervical cancer cell line SiHa and HPV18 positive cervical cancer cell line HeLa,then qRT-PCR was used to detect the expression of miR-543 and the expression of CHMP4C was detected by Western blotting,thus clarifying the relationship between HPV E6 and miR-543 and CHMP4C.5.The dual-luciferase reporter assay was used to verify whether there was a connection site between CHMP4C and miR-543.Results:1.The results of immunohistochemistry showed that the level of CHMP4C expression in cervical cancer tissues was higher than that in normal tissues,and the level of CHMP4C was related to the age of the patients(P<0.05).2.Down-regulation of CHMP4C reduced proliferation,migration and invasion of cervical cancer cells,and increased cell apoptosis(P<0.05).3.Western blotting showed that silencing CHMP4C reduced the expression of apoptotic regulatory protein bcl2 and bcl-xl.4.After knocking down E6,miR-543 expression was elevated,while CHMP4C expression decreased,indicating that E6 can negatively regulate miR-543 expression and positively regulate CHMP4C expression.5.The result of Dual Luciferase Report showed that there were connection sites between miR-543 and CHMP4C(P<0.05).Conclusion:HPV E6 may regulate the expression of CHMP4C through miR-543 to promote the tumorigenesis and development of cervical cancer.