Expression,Clinical Significance,Function And Mechanism Of TRNA-Derived Fragment 3019a In Gastric Cancer

Objective:Gastric cancer(GC)is one of the most common human cancer types with an increasing global burden and the second leading cause of cancer-related deaths worldwide.Since it is often diagnosed at an advanced stage,treatment with current surgical,chemical or radiological therapies often cannot improve patient prognosis,resulting in a poor survival rate.More effective and accurate diagnostic techniques would benefit GC patients by identifying GC at earlier stages.However,current conventional diagnostic methods have limitations in specificity and sensitivity.Therefore,it is imperative to identify novel,stable,and effective biomarkers to better diagnose and screen GC patients at an early stage.Many studies demonstrate that tRNA fragments are not products of random tRNA degradation,rather they are generated by multiple pre-tRNAs and mature tRNAs undergoing endonucleolytic cleavage with different ribonucleases.Moreover,increasing evidence has demonstrated that t RNA fragments exert important effects on various human cancer types.However,it is still unknown in GC.Methods:For the first section,we tested tRNA-derived fragment 3019a(tRF-3019a)levels in HGC-27,MGC-803,AGS,MKN-45,SNU-16 and GES-1 using real-time quantitative reverse transcription-polymerase chain reaction(qRT-PCR).Then,tRF-3019 a levels in 112 paired GC tissues and adjacent non-tumorous tissues from patients with GC were detected by qRT-PCR.Furthermore,we determined the clinicopathological factors and prognostic information of patients to explore its clinical significance.A receiver operating characteristic curve was generated to evaluate the diagnostic value of tRF-3019 a.For the second section,its biological effects were investigated by cell proliferation assay,cell cycle analysis,cell apoptosis analysis,cell migration,and invasion assay and western blot analysis.The downstream target genes of tRF-3019 a were predicted using bioinformatics and databases,and the downstream target genes were screened by the combination of GO function enrichment and KEGG.Luciferase reporter assaywas used to verify the direct binding relationship between tRF-3019 a and downstream target gene FBXO47(F-box protein 47).The function of FBXO47 was investigated in GC.We sought to investigate whether tRF-3019 a affects GC biological function by targeting FBXO47 using a “rescue” experiment.Results:tRF-3019 a was found to be significantly upregulated in GC tissues and cell lines and was not related to the clinicopathological factor and survival time of the patients.The area under the ROC curve was 0.689 for all GC patients(P<0.001).Loss-of-function and gain-of-function studies,using inhibitors and mimics of tRF-3019 a respectively,revealed that tRF-3019 a enhanced migration and invasion in HGC27 and MGC803 cells.However,we found no significant change in cell cycle or apoptosis when tRF-3019 a was knocked down.FBXO47 was determined to be the downstream directly regulated target gene of tRF-3019 a.Silencing FBXO47 can enhance the proliferation,migration,and invasion of GC cells.Mechanism investigation implicated FBXO47 as a downstream target of tRF-3019 a binding through which it enhanced tumor migration and invasion phenotypes.Conclusion:Our findings reveal that tRF-3019 a is upregulated in GC tissues and cell lines.tRF-3019 a would become a novel biomarker for the diagnosis of GC.tRF-3019 a may affect GC cell migration and invasion by targeting FBXO47.