NUDT21 Regulates SGPP2 To Promote The Malignant Progression Of Gastric Cancer

BackgroundGastric cancer is one of the most common malignancies not only in our country but also worldwide.In China,gastric cancer accounts for about 20% of all deaths from malignant tumors.At present,the first choice of treatment for gastric cancer is surgical resection,followed by post-operative radiotherapy.However,the recurrence rate of gastric cancer remains high and the prognosis of patients is poor.Further research into the molecular mechanisms underlying gastric cancer tumourigenesis,proliferation and metastasis is urgently needed to improve clinical outcomes.NUDT21(naked sea medium hydrolase 21)is an RNA-binding protein complex that can act as a post-transcriptional regulator to establish a direct link between alternative polyadenylation and chromatin signaling.NUDT21 functions with tissue specificity and plays an oncogenic role in human pancreatic ductal adenocarcinoma cells and leukemia cells,but in our previous study,we found that NUDT21 can inhibit proliferation and metastasis of human hepatocellular carcinoma and breast cancer cells.However,the regulatory role and regulatory mechanism of NUDT21 in human gastric cancer cells need to be further explored.AimInvestigating the expression pattern and mechanism of action of NUDT21 in gastric cancer.Methods1)The expression level of NUDT21 was explored by immunohistochemistry and qPCR experiments in clinical samples collected from gastric cancer patients.It was further validated by qPCR and protein blotting experiments in GES-1(gastric normal epithelial cells)and AGS,BGC-823,SGC-7901,HGC-27,MKN-28(five gastric cancer cell lines).Cancer and paraneoplastic tissues from 70 gastric cancer patients were collected to analyze the association between NUDT21 expression levels and clinicopathological features of gastric cancer patients.2)NUDT21 overexpression and knockdown transient cell lines were constructed by plasmid/oligonucleotide transfection in gastric cancer cell lines MKN-28 and BGC-823,and the knockdown and overexpression efficiency of NUDT21 was verified by qPCR and protein blotting assay;the effects of NUDT21 on proliferation,migration and invasion of gastric cancer cells were investigated in vitro by MTT assay,colony formation assay,EdU proliferation assay and Transwell assay.3)MKN-28 gastric cancer cells with low NUDT21 expression level were selected to establish a NUDT21 overexpression transient cell line,and subcutaneous tumorigenesis assay were performed in nude mice;BGC-823 gastric cancer cells with high NUDT21 expression level were selected to establish a NUDT21 knockdown transient cell line,and tail vein injection assay were conducted in nude mice to study the effect of NUDT21 expression on the progression of gastric carcinogenesis in vivo.4)The downstream gene SGPP2,which was significantly regulated by NUDT21,was screened by high-throughput sequencing,and the effect of altered NUDT21 expression level on SGPP2 was further verified by qPCR and western blot assays,and the correlation between NUDT21 and SGPP2 expression in 70 gastric cancer tissues was analyzed by Pearson correlation analysis.5)NUDT21 knockdown and SGPP2 overexpression plasmids were cotransfected in BGC-823 cells,and NUDT21 overexpression plasmids and si RNA targeting SGPP2 were cotransfected in MKN-28 cells to investigate the regulation of SGPP2 on the role of NUDT21 in gastric cancer by MTT assay,colony formation assay,EdU proliferation assay and Transwell assay.Results1)NUDT21 was expressed at much higher levels in human gastric cancer cells / tissues compared to normal gastric cells / tissues.Tumor tissues from patients with distal tumor metastasis showed elevated levels of NUDT21 compared to tumor tissues from patients without distal tumor metastasis.High levels of NUDT21 expression were associated with tumor size,lymph node metastasis and clinical stage in gastric cancer patients,and patients with high NUDT21 expression had poorer OS rates compared to patients with low NUDT21 expression.2)MTT assay,colony formation assay,EdU assay,cell migration assay and cell invasion assay demonstrated that sh RNA-mediated knockdown of NUDT21 significantly reduced cell viability and inhibited migration and invasion of gastric cancer cells.Overexpression of NUDT21 promoted cell proliferation and metastasis of gastric cancer cells.3)Overexpression of NUDT21 promoted the growth and proliferation of gastric cancer tumor cells,and knockdown of NUDT21 inhibited lung metastasis of gastric cancer cells in subcutaneous tumorigenesis assay and lung metastasis assay in nude mice.4)For the downstream mechanism of NUDT21 regulation,SGPP2 is positively regulated by NUDT21 in gastric cancer cells.5)Overexpression of SGPP2 rescued the downregulation in cell proliferation and metastasis induced by NUDT21 knockdown in BGC-823 cells;knockdown of SGPP2 rescued the upregulation in cell proliferation and metastasis induced by NUDT21 overexpression in MKN-28 cells.ConclusionGastric cancer tissues express high levels of NUDT21.High levels of NUDT21 are associated with clinicopathological features and poor survival in gastric cancer patients.SGPP2 is positively regulated by NUDT21 and mediates the carcinogenic effects of NUDT21 in gastric cancer cells.