The Mechanism Of SHARPIN In Regulating P53 Ubiquitination And Degradation In Breast Cancer

Background: Breast cancer is the most common malignancy in women,and the second cause of death in women.In our country,the number of new cases of breast cancer accounts for 12.2% of all over the world,while the number of deaths accounted for 9.6% of the world.Although the prognosis of patients with early breast cancer is better,but the prognosis of patients with advanced breast cancer is still poor,so the need for accurate diagnosis of tumor markers and specific targets and drugs.By analysis of TCGA public available database(https://tcga-data.nci.nih.gov/),we observe that SHARPIN m RNA level is higher compared with normal breast tissue.In addition,there are many female patients who have breast cancer family history often exist p53,PTEN,BRCA1 / 2 gene mutation.It is important to identify the effects of these genes in breast cancer and to find new drug targets and inhibit the development and progression of breast cancer.Objective: To explore the mechanism of SHARPIN protein and p53 protein in breast cancer cell lines,so as to provide new ideas for new drug targets treatment in breast cancer.Methods: 1.Using the TCGA and Kmplot database to clarify the expression of SHARPIN in human breast cancer tissue and the relationship between SHARPIN expression and prognosis of patients.Using genome-wide RNA sequencing to investigate whether SHARPIN affects p53 signaling pathway.2.The effect of SHARPIN in the proliferation of breast cancer cell lines was confirmed by WST-1 cell proliferation assay and EDU experiment.3.The relationship of SHARPIN and p53 signaling pathway is confirmed by q RT-PCR and Western Blot experiments.4.The results showed that SHARPIN affected the post-translational modification of p53 protein by q RT-PCR ?Western Blot?MG132 and CHX experiments.5.The intermediate bridge of SHARPIN indirect control of p53 protein stability was found by Co-IP,MG132 and CHX experiments.6.The relationship between SHARPIN and MDM2 protein stability was confirmed by CHX and MG132 experiments.Results: 1.SHARPIN and p53 wild-type breast cancer patients have a certain relationship between the poor prognosis.RNA-seq results demonstrate that silencing of SHARPIN affects the expression of the p53 target gene.2.SHARPIN silencing can decrease breast cancer cell proliferation ablity.3.SHARPIN silencing can upregulate p53 signaling pathway activity.4.SHARPIN can affect post-translational modification of p53 protein.5.SHARPIN moldulate p53 protein stability in MDM2 dependent manner.6.SHARPIN can increase MDM2 protein stability.Conclusions: In breast cancer cells,SHARPIN depend MDM2 to regulate ubiquitination and degradation of p53 protein,and then affect the stability of p53 protein.