The Function And Upstream Regulatory Mechanism Of SRSF10 In Promoting Malignant Phenotype Of Liver Cancer Cells

Background: The global morbidity and mortality of malignant tumors are increasing rapidly,and it has become one of the most serious public health problems in the world.In my country,primary liver cancer is currently the fourth most common malignant tumor and the second leading cause of tumor death,and the overall morbidity and mortality in the population are still showing an upward trend.Because most patients with liver cancer have no obvious symptoms in the early stage,the diagnosis is difficult,and the treatment methods for the advanced stage patients are very limited,the effect is not good,and the prognosis is very poor.Therefore,there is an urgent need for new targets that can be applied to the diagnosis and treatment of liver cancer.Liver cancer is caused by a complex combination of genetic and epigenetic modification alterations.In recent years,a large number of studies have shown that the epigenetic phenomenon m6 A methylation is closely related to the occurrence and development of malignant tumors.The previous study of our group found that the m6 A modification key gene SRSF10 may be highly expressed in liver cancer.However,the role and molecular mechanism of SRSF10 in the occurrence and development of hepatocellular carcinoma have not been fully understood.Therefore,this topic strives to provide new targets and new ideas for the clinical diagnosis and treatment of liver cancer.Methods:1.Four hepatocellular carcinoma datasets,GSE87630,GSE46408,GSE84402,and GSE101685,were obtained from the GEO database and comprehensively analyzed,and the data on the expression level,clinical stage and survival time of SRSF10 were obtained through the UALCAN database,GEPIA database,and TCGA;2.10 fresh tissues from clinical specimens of liver cancer and 10 fresh tissues adjacent to the control cancer were collected.Western Blot and immunohistochemistry were used to detect the expression of SRSF10 protein.Nine liver cancer cell lines Huh7,Hep G2,SMMC-7721,Hep3 B,BEL-7402,BEL-7404,MHCC97 H,MHCC97L,MHCC-LM39 and 1normal liver cell line L02 were detected by Western Blot and q RT-PCR to detect the protein and m RNA expression of SRSF10;3.Hep G2 and Hep3 B hepatoma cells were transfected with blank plasmid,SRSF10-OE,SRSF10-KD1,SRSF10-KD2,and the expression of SRSF10 protein and m RNA was detected by Western Blot and q RT-PCR;4.the CCK-8 proliferation assay,Edu fluorescence proliferation assay and Transwell assay were performed to explore the biological role of SRSF10 in malignant phenotypes such as proliferation,cell cycle and migration of liver cancer cells;5.NCBI,JASPAR,GEPIA,UALCAN and other databases were used to search and screen the upstream transcription factor TLX1 of SRSF10 gene;Taking Hep3 B and Hep G2 liver cancer cell lines as research objects,Ch IP assay was used to detect the binding ability of SRSF10 and TLX1,and Western Blot and q RT-PCR experiments were used to confirm that TLX1 regulates the expression of SRSF10;6.The Hep3 B liver cancer cell line was transfected with blank plasmid,si-TLX1,si-TLX1+OE-TLX1,and the CCK-8 proliferation assay and flow cytometry were performed to verify that TLX1 regulates the function of SRSF10 gene in liver cancer.Results:1.The results of bioinformatics analysis showed that SRSF10 was highly expressed in liver cancer.SRSF10 is up-regulated in clinically advanced hepatocellular carcinoma(P<0.01).The expression level of SRSF10 was negatively correlated with OS(P<0.01)and DFS(P<0.05)in patients with HCC.2.Compared with adjacent tissues,SRSF10 protein showed high expression in liver cancer tissue samples.Compared with the normal liver cell line L02,SRSF10 was significantly elevated in most liver cancer cell lines.3.Compared with the control group,the protein and m RNA expression of SRSF10 was significantly decreased after knockdown,while the protein and m RNA expression of SRSF10 was significantly increased after overexpression of SRSF10.4.Compared with the blank control group,overexpression of SRSF10 can promote the proliferation and invasion of hepatocellular carcinoma cells;SRSF10 can inhibit the proliferation and invasion of liver cancer cells.5.The results of Ch IP experiment showed that the binding ability of transcription factors TLX1 and SRSF10 was enhanced after high expression of TLX1,and the binding ability of TLX1 and SRSF10 was weakened after TLX1 knockdown.The results indicated that the transcription factor TLX1 directly and positively regulates SRSF10.Western Blot and q RT-PCR experiments showed that knockdown of TLX1 could significantly down-regulate the expression of SRSF10.6.The functional recovery experiment showed that the apoptosis rate of TLX1 knockdown was higher than that of the control group,and the cell proliferation rate was decreased.On this basis,overexpression of SRSF10 could reverse this result.Conclusion: SRSF10 is highly expressed in HCC and negatively correlated with the prognosis of HCC patients.SRSF10 can promote the proliferation and invasion of liver cancer cells.The transcription factor TLX1 can promote the expression of SRSF10 through transcription,thereby promoting the proliferation of liver cancer cells and inhibiting apoptosis.