The Molecular Mechanism In Circular RNA CircSMAD2 Affecting Proliferation Of Breast Cancer Cells

Circular RNA belongs to the long-non-coding RNA family.Circular RNA is formed by "back splicing" in the process of RNA processing,which is formed by the precursor mRNA through the linkage between head and tail.Due to the absence of the 3' poly A structure and the cap structure at the 5' end,circular RNAs are not recognized by RNA exonuclease and can be stably present in cells for a long time.Early high-throughput sequencing studies have shown that circular RNA is ubiquitous in animal cells.In recent years,many studies have also shown that circular RNA is closely related to the occurrence and development of diseases,but many mechanisms are still unclear.Breast cancer is a kind of high-risk disease in the female population,and its morbidity and mortality have been ranked first in all ages.Molecular targets related to breast cancer treatment have been a hot topic in the world,but most of the research focuses on oncogenes and tumor suppressor genes.In this paper,this research has conducted a deep analysis on the effect of circSMAD2 that formed by Smad2,which is one of the important factors in the TGF? signaling pathway.The specific research contents are as follows:(1)Firstly,it was found that there was a correlation between various cancer and circular RNA through Meta-analysis: By searching and screening the literatures that is related circular RNA in the common databases(PubMed,Embase,Web of Science).We identified 19 sets of clinical data from 1752 patients of 17 research papers with complete clinical data for human cancer,and Meta-analyzed to calculate the sensitivity(sensitivity,SEN): 0.72,specificity(specificity,SPE): 0.74,likelihood ratio(LR): 2.80 and 0.38,diagnostic odds ratio(DOR): 7.00 and area under curve(AUC): 0.79 of the expression of circRNA.The results indicate that circRNA has potential value as a marker for clinical detection of tumors.(2)Then,we examined the effect of circSMAD2 on breast cancer cells: We successfully constructed the overexpression vector of circular RNA circSMAD2 and performed it by semi-qPCR.It was transfected into breast cancer cells,and the function of circSMAD2 was verified by cell function experiments: MTT assay and colony formation assay demonstrated that circSMAD2 can inhibit the proliferation of breast cancer cells;then the cell cycle was detected by flow cytometry and it was found that circSMAD2 arrests cells in the S phase of the cell cycle and inhibits the proliferation of breast cancer cells.(3)Finally,the molecular mechanism of the effect of circSMAD2 on cell cycle was analyzed: We overexpressed circSMAD2 in the breast cancer cells,and the expression levels of cell cycle associated proteins and upstream proteins p53 were detected by Western blot.circSMAD2 can promote the expression of p53 and increase the expression of p21,but has no effect on the transcription levels of cyclin and p53,and the level of ubiquitination of p53 is reduced.It can be preliminarily proved that the inhibition of the cell cycle by the circular RNA circSMAD2 is caused by inhibiting the degradation of p53 protein,causing cell cycle changes,and arresting breast cancer cells in the S phase of the cell cycle.