The Pseudogene Derived Long Noncoding RNA DUXAP8 Promotes Gastric Cancer Cell Proliferation And Migration Via Epigenetically Silencing PLEKHO1 Expression

BackgroundGastric cancer is one of the most common malignant gastrointestinal tumors in China,the morbidity and mortality of malignant tumors accounted for 2nd and 3rd.The progression and development of gastric cancer physiological process is regulated by a variety of factors.Recently,evidence indicates that pseudogenes is involved in a wide range of cell biological processes.Pseudogenes are highly similar to encoding proteins genes,but cannot express of gene inside the cell.Moreover,some pseudogenes transcribed non-coding RNAs more than 200 nt in length cloud be included in IncRNA classification.Dysregulated IncRNAs expression was associated with tumor generation and development,and exercise the function of oncogenes or tumor suppressor genes.The pseudogene derived IncRNA DUXAP8 biological function and potential mechanism in gastric cancer cells is still unclear.ObjectiveThe aim of this study is to detect DUXAP8 expression in gastric cancer and to evaluate the role and potential mechanisms of DUXAP8 in the progression and development of gastric cancer.Methods(1)We analyzed the expression levels of DUXAP8 in human gastric cancer tissues by using raw microarray data downloaded from GEO;qRT-PCR was used to testify the expression of DUXAP8 in gastric cancer tissues;We assess the correlation between DUXAP8 expression and clinicopathological features.(2)MTT and clone formation assays were used to evaluate the viability of gastric cancer cells;Flow-cytometric analysis was performed to characterize the level of apoptosis and cell cycle regulation;Transwell experiment weer performed to evaluate gastric cancer cell migration.(3)RNA transcriptome sequencing weer used to find the DUXAP8-associated pathway.(4)RIP and ChIP assays were performed to evaluate the potential mechanism that DUXAP8 how to regulate the downstream target gene.(5)Build PLEKHO1 expression plasmid,and by saving experiment DUXAP8 could regulate the expression of downstream target gene.(6)Nude mouse transplantation tumor model experiment was performed to observe the effect of gastric cancer cell transfected with si-DUXAP8 on tumor ability in vivo.Results(1)The GEO and qRT-PCR results showed that the DUXAP8 expression levels were upregulated in gastric cancerous tissues compared with noncancerous tissues.(2)High DUXAP8 expression in gastric cancer was significantly correlated with tumor size,advanced TNM stage,lymph node metastasis and prognosis of patients.(3)Knockdown DUXAP8 inhibited gastric cancer cell proliferation,metastasis,and promoted cell apoptosis.(4)Analysis of the RNA transcriptome sequencing data from triplicate samples revealed that a common set of 133 mRNAs exhibited increased expression in DUXAP8-depleted cells,while 315 mRNAs were downregulated.qRT-PCR detection further found that PLEKHOl might be DUXAP8 downstream target gene.(5)RIP and ChIP assays showed that DUXAP8 may epigenetically inhibit PLEKHO1 by binding to PRC2.(6)Rescuing experiment results show that raising expression of PLEKHO1 can save the effect of DUXAP8 promote proliferation.(7)The results of nude mouse transplantation tumor model experiment showed that the inhibition of DUXAP8 expression can reduce tumor ability in vivo.ConclusionThe study firstly demonstrated that DUXAP8 cloud promote gastric cancer cell proliferation and migration via epigenetically silencing PLEKHO1 expression.our results provide a new perspective that DUXAP8 could be a novel target for the early diagnosis and treatment of gastric cancer.