Study On The Mechanism Of Syntenin Affecting The Invasion And Migration Of Pancreatic Cancer By Regulating YAP1

Melanoma differentiation related gene mda-9(also known as syntenin-1;SDCBP(polyligand glycan binding protein)is a transfer-promoting gene,which was initially found in human metastatic melanoma cells [1,2].In previous studies,mda-9/Syntenin was found to be significantly up-regulated in melanoma,uveal melanoma,glioblastoma multiforme,breast cancer,small cell lung cancer,urinary epithelial tumor,head and neck cancer,prostate cancer,etc.,and was significantly correlated with prognosis [3-10].The high metastasis of pancreatic cancer is an important clinical problem that restricts its treatment.Therefore,as a potential oncogene,Syntenin is expected to become an important biomarker in tumor diagnosis and treatment.However,the specific mechanism of Syntenin in the development of pancreatic cancer has not been reported.Found in this study,Syntenin in pancreatic cancer(PDACs)expression levels were significantly higher than those in the tissue adjacent to carcinoma,and the expression level Syntenin and patients with tumor size,lymph node metastasis and histological grading of remarkable relevance,more important is Syntenin expression level and the overall survival,relapse-free survival in patients with a significantly negative correlation between.Methods 1.IHC of Syntenin was performed in 104 cases of pancreatic cancer and para-cancer paired tissue chips,and survival statistical analysis was performed.8 cases of pancreatic cancer and para-cancer paired tissue samples were collected for verification by western-blot and QPCR.2.The crisper technology was used to construct the overexpression and knockout stability system of Syntenin,and experiments were conducted on transwell,scratch,western-blot detection of EMT index,edu and RTCA.3.The in-situ model of pancreatic cancer in mice and the liver metastasis model of pancreatic cancer injected into the spleen were established.4.IHC of syntenin and YAP1 was performed by continuous chip sections of pancreatic cancer,and the correlation between the two was statistically analyzed.Theexpressions of SDCBP and YAP1 were detected by western blot and immunofluorescence.5.Blocking YAP1 for scratching,transwell,edu and western-blot detection of EMT indicators,in vivo imaging technology and dissection of mice to detect and observe tumor size and distant metastasis.6.Add CHX to control and stable system,extract proteins at 0h,30 min,2h,3h,4h and 6h,detect the expression of YAP1 by western-blot,instantaneously transfer the plasmid of cmyc-yap1 in control and stable system,and repeat the above steps.7.Add MG132 to control and stable lines for 12 h,add CHX,extract proteins at 0h,30 min,2h,3h,4h and 6h,and detect the expression of YAP1 by western-blot.8.Exogenous and endogenous ubiquitin IP experiments of cmyc-yap1 and YAP1 were conducted in 293 T and bxpc-3 cell lines,respectively.9.IP experiments of Syntenin and YAP1 were conducted in 293 T and pancreatic cancer cells,respectively.Truncated plasmids of Syntenin and YAP1 were constructed and IP experiments were carried out in 293 T.10.The plasmid of flag-yap1 /△TAD was transiently transferred in control and stable system,and CHX was added after 48 h.Proteins were extracted at 0h,30 min,2h,3h,4h and 6h,and the expression of flag-yap1 /△TAD was detected by western-blot.11.Transient overexpression of YAP1 plasmid and small interference of YAP1 in pancreatic cancer cells.The expression level of Syntenin was detected by Western-blot and QPCR.TEAD and YAP1 co-transcriptional TEAD transcriptional regulation of Syntenin by CHIP and double luciferase experiments were conducted.12.Drugs that can affect the stability of Syntenin protein in vitro were selected from small molecular synthesis library.The expression of Syntenin,the changes of EMT and the transwell assay were detected by western-blot.Results and Conlusions 1.The expression of Syntenin in cancer was higher than that in paracancer tissues,and it was negatively correlated with the prognosis.2.Syntenin can promote theinvasion,migration,EMT and proliferation of pancreatic cancer cells.3.There is an obvious positive correlation between Syntenin and YAP1.Syntenin promotes invasion,migration,metastasis,and proliferation of pancreatic cancer through YAP1.4.Syntenin can inhibit the ubiquitination of YAP1 and further inhibit the degradation of the proteasome pathway of YAP1.5.YAP1 was combined with the ha-pdz1 and ha-pdz2 structural domains of Syntenin through flag-tad structural domain.6.YAP1 was positively correlated with Syntenin at m RNA and protein levels.TEAD1 can directly regulate the expression of Syntenin by transcriptional regulation,and YAP1 can enhance the expression of Syntenin by transcriptional regulation of TEAD1.7.Rabeplazole sodium can significantly inhibit the expression of Syntenin,EMT and invasion and migration of pancreatic cancer cells.