Function Of Enhancer RNA Induced Expression Change Of PP1A On Proliferation And Metastasis Of Hormone Receptor Positive Breast Cancer

Background In 2020,the world cancer statistics released by International Agency for Research on Cancer of WHO showed that breast cancer has surpassed lung cancer as the world’s largest malignant tumor,with the incidence rate of 11.7% of all new cancer cases.In 2020,the number of newly diagnosed cancer cases in China was 2,090,000,of which 420,000 cases were newly diagnosed breast cancers,ranking the first place in malignant tumors for female.Breast cancer brings serious physical and psychological burden to patients.Prevention,early diagnosis and treatment of breast cancer are particularly important.Breast cancer is a well-defined malignant tumor.According to the status of hormone receptor and Ki67 expression,breast cancer can be divided into Luminal A type breast cancer,Luminal B type breast cancer,HER2 overexpression breast cancer and triple negative breast cancer.There are differences in prognosis,drug responsiveness,recurrence rate and metastasis rate of breast cancer with different molecular type.Hormone receptor positive breast cancer is sensitive to endocrine therapy,but there are also problems of recurrence and metastasis.Hippo-YAP1 signaling pathway plays an important role in regulating tissue and organ development,tumor development and so on.The role of Hippo-YAP1 signaling pathway in hormone receptor positive breast cancer has been confirmed by many studies.The analysis of protein-protein interaction network showed that PP1 A protein is a potential interaction partner of YAP1.PP1 A is the α subunits of protein phosphatase 1,which is involved in the phosphorylation modification of specific proteins.However,the biological functions and molecular mechanisms of PP1 A in hormone receptor positive breast cancer remained unclear.Therefore,the purpose of this study was to investigate the role of PP1 A in the proliferation and metastasis of hormone receptor positive breast cancer and the specific upstream and downstream molecular mechanisms.Purposes(1)To explore the expression of PP1 A in hormone receptor positive breast cancer.(2)To explore the correlation between PP1 A expression and clinical characteristics such as hormone receptor expression status of breast cancer patients.(3)To explore the biological function of PP1 A in hormone receptor positive breast cancer.(4)To explore the regulation of PP1 A on Hippo-YAP1 signal pathway.(5)To explore regulation function of enhancer RNA_LINC02574 on expression of PP1 A.Methods(1)Real time fluorescence quantitative PCR and Western blot were used to detect the expression of PP1 A m RNA in breast cancer tissues and breast cancer cell lines on RNA and protein level.The correlation between PP1 A and breast cancer molecular type,axillary lymph node metastasis and other clinical characteristics were analyzed.(2)Small interfering RNA specifically targeted on PP1 A mRNA was used to transfect breast cancer cells to achieve PP1 A knockdown,and PP1 A lentivirus plasmid was used to transfect breast cancer cells to achieve overexpression of PP1 A.(3)The effects of PP1 A on proliferation and metastasis of breast cancer cells were explored by MTT cell proliferation assay,plate clone formation assay,wound healing assay,Transwell assay and tumor formation in nude mice.(4)Western blot and protein immuno-coprecipitation were used to explore the key downstream signal pathway of PP1 A.(5)The key transcription factors of PP1 A were analyzed by Dual Luciferase Reporter Gene Assay.(6)Interaction between enhancer RNA_LINC02754 and PP1 A was explored by RNA immune-coprecipitation and RNA pulldown assay.Results(1)Compared with adjacent normal tissues,PP1 A expression is increased in breast cancer tissues.On comparison with MCF10 A cells,the expression of PP1 A increased in MCF7,BT474,SKBR3 and MDA-MB-231 cells,especially in MCF7 and BT474 cells.(2)After PP1 A knockdown,the proliferation and migration ability of hormone receptor positive breast cancer cells MCF7 and BT474 were decreased.After overexpression of PP1 A,the proliferation and migration ability of hormone receptor positive breast cancer cells MCF7 and BT474 were increased.The results of tumorigenesis in nude mice showed that PP1 A could promote the proliferation of breast cancer cells in vivo.(3)PP1A interacts with transcription cofactor YAP1,inhibits the phosphorylation of YAP1,and then activates the expression of downstream oncogenes of YAP1.(4)β-estradiol can induce hormone receptor positive breast cancer cell lines MCF7 and BT474 expression enhancers RNA_LINC02754.The enhancer RNA_LINC02754 recruits transcription factor E2F1 to bind to the promoter region of PP1 A gene and activates the transcription of PP1 A.ConclusionsPP1A is upregulated hormone receptor positive in breast cancer cells.In terms of biological function,PP1 A can promote the proliferation and metastasis of hormone receptor positive breast cancer cells.In terms of molecular mechanism,estrogen induced enhancer RNA_LINC02754 recruits transcription factor E2F1 and binds to the promoter region of PP1 A gene to promote PP1 A transcription.PP1 A inhibits the phosphorylation of YAP1 by interacting with YAP1,a key molecule of Hippo-YAP1 signaling pathway.Dephosphorylated YAP1 is activated,shifts from cytoplasm to nucleus,and promotes the expression of downstream oncogenes.This research describes the biological functions and specific molecular mechanisms of PP1 A in hormone receptor positive breast cancer,which may provide a new perspective for the diagnosis and treatment of hormone receptor positive breast cancer.