| ObjectiveCervical cancer(CC) which frequently happens in women all around the world is the fourth reason of woman-related mortality. Although some different approaches to the treatment of cancer have decreased the incidence and mortality, such as prophylactic HPV vaccines and traditional pap-smear screening, a lot of women died of resisting to regular treatment and late diagnosis every year. MicroRNAs(miRNAs) consist of a sequence of small non-coding RNAs whose function are involved in gene expression and cellular pathways. They could participate in the transformation from precancerous lesion to invasion and metastasis, thus promoting tumour malignancy. In fact, a great deal of evidence has demonstrated that miRNAs play evident role in normal metabolism and cellular homeostasis. Therefore, deregulation of these molecules is related with the incidence of diseases.?-1,4-Galactosyltransferase III(B4GALT3) is an enzyme responsible for the generation of poly-N-acetyllactosamine and is involved in tumorigenesis. However, B4GALT3-dysregulation and its role in cervical cancer cells are unknown. Our aim is to investigate the influence of B4GALT3 on malignant phenotype as well as the upstream and downstream mechanisms underlying the phenomenon, thus clarifying the effects of B4GALT3 on ceivical carcinogenesis. MethodsFirst, we detected the expressions of B4GALT3 in cervical cancer tissues and adjacent non-tumor tissues using RT-qPCR. Then wound healing assays, transwell migration/matrigel assays and Western blot assays were performed to investigate the migration and invasion ability as well as the process of EMT. Furthermore, miRNA which could target the 3'UTR of B4GALT3 was predicted by the bioinformatics analysis and levels of B4GALT3 mRNA and protein regulated by miR-27 a were detected by RT-qPCR and Western blot. At the same time, EGFP and luciferase reporter system were performed to further demonstrate that B4GALT3 was directly targeted by miR-27 a. The mechanism under which B4GALT3 was upregulated by miR-27 a was investigated by mRNA and protein degradation assays. We also investigated the malignant phenotypes induced by miR-27 a. At the same time, B4GALT3 was knocked down in miR-27a-overexpressing cells and under another circumstance miR-27 a was blocked in B4GALT3-overexpressing cells to investigate the malignant phenotype changes. Finally, we performed protein degradation assays as above to detect the stability of ?1-integrin induced by B4GALT3. Results B4GALT3 was upregulated in cervical cancer tissues compared to adjacent non-tumor tissues. B4GALT3-overexpression promoted, whereas B4GALT3-knockdown suppressed the cellular migration, invasion and EMT of HeLa and C33 A cervical cancer cells. To explore the mechanism of dysregulation, B4GALT3 was predicted to be a target of miR-27 a. EGFP and pGL3-promoter reporter assays showed that miR-27 a bound to B4GALT3 3'UTR region but enhanced its expression. RT-qPCR showed that miR-27 a also overexpressed and presented positive correlation with B4GALT3-expression in cervical cancer tissues. miR-27a-overexpression promoted, but blocking-miR-27 a repressed these malignancies in HeLa and C33 A cells. Furthermore, shR-B4GALT3 counteracted the promotion of malignancies induced by miR-27 a, suggesting that miR-27 a upregulates B4GALT3 to enhance tumorigenic activities. In addition, we found that B4GALT3 significantly enhances ?1-integrin stability, thus mediating promotion of B4GALT3 on malignancy in cervical cancer cells. Altogether, our findings evidenced that B4GALT3-upregulated by miR-27 a contributes to the tumorigenic activities by ?1-integrin pathway and might provide potential biomarkers for cervical cancer. ConclusionsB4GALT3 is up regulated by miR-27 a and causes malignant phenotype thro?gh enhancing the stability of ?1-integrin. |
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