MYO16-AS1 Inhibits The Migration And Invasion Of Lung Adenocarcinoma By Regulating HK2 MRNA Stability

BackgroundLung cancer is a malignant tumor with the highest morbidity and mortality in China.lung cancer is mainly divided into Small cell lung cancer(SCLC)and Non-small cell Lung cancer(NSCLC)according to pathological types,including Lung Adenocarcinoma(SCLC),LUAD,as the pathological type with the highest proportion in NSCLC,accounts for about 35%of all lung cancers,and has an increasing trend year by year.In recent years,with the development of clinical testing techniques and treatment methods,the 5-year survival rate of early LUAD patients has reached more than 90%.However,for patients with mid-and late-stage LUAD,the prognosis is often poor due to the extensive local invasion and distant metastasis of LUAD.Therefore,it is important to explore the molecular mechanism related to LUAD infiltration and migration to improve the prognosis of LUAD patients.Long-stranded non-coding RNA(LncRNA)is a non-coding RNA with a length greater than 200 nucleotides.Lncrnas participate in cellular physiological or pathological activities by interacting with RNA-binding proteins(RBPS)in cells,and their main functions include:chromatin remodeling,adsorption,participation in protein complex formation,transcriptional activation,transcriptional inhibition,translation inhibition,shear regulation,post-translational degradation of mRNA,etc.Recent studies suggest that lncrnas play an important role in the occurrence and development of tumors.However,the molecular mechanism of lncrnas involved in the occurrence and development of LUAD remains unclear,so further exploration is needed.Objectives1.Screening and verification of LncRNA that play a key role in the occurrence and development of LUAD.2.To explore the role and molecular mechanism of MYO16-AS1 in LUAD infiltration and migration.Methods1.RNA-seq analysis was performed on 7 groups of tissue samples collected clinically from LUAD patients,and lncrnas with different expressions in LUAD tumor cells were screened out by comparing the data in GTEx database and TCGA database.2.MYO16-AS1 overexpression and knockdown cell lines were constructed,and the effects of MYO16-AS1 on the migration and invasion ability of LUAD were detected by Transwell assay.The effect of MYO16-AS1 overexpression on tumor formation in nude mice was detected by subcutaneous tumor transplantation experiment.3.Relevant proteins were extracted by RNA pulldown,Coomasil bright blue staining was performed on protein glue,potential binding RBP with MYO16-AS1 was screened out by comparing with public databases,and the binding relationship between MYO16-AS1 and IGF2BP3 was proved by RNA pulldown and RIP experiments.The intracellular localization of MYO16-AS1 and IGF2BP3 was determined by fluorescence in situ hybridization(FISH)and immunofluorescence assay.4.The intersection of IGF2BP3 iCLIP results and related down-regulated genes in RNA-seq data was used to analyze downstream target genes.The influence of MYO16-AS1/IGF2BP3 complex on HK2 mRNA expression level was detected by Western blot and RT-qPCR,and the influence of overexpression of HK2 on intracellular glucose content was detected by glucose content determination.5.The relationship between IGF2BP3 and HK2mRNA was predicted by Starbase database,and the spatial structure relationship between MYO16-AS1/IGF2BP3 and IGF2BP3/HK2mRNA was predicted by HDock database.Results1.The expression of MYO16-AS1 was significantly down-regulated in LUAD tissues and cell lines.2.In vitro experiments,overexpression of MYO16-AS1 inhibited the invasion and migration ability of LUAD tissue cells.In vivo,overexpression of MYO16-AS1 inhibited the tumorigenic ability of LUAD.3.MYO16-AS1 directly aggregates with IGF2BP3 in the cytoplasm.4.MYO16-AS1/IGF2BP3 complex reduced the stability of HK2 mRNA and inhibited glucose metabolism in cells,thus inhibiting the invasion and migration of LUAD.5.MYO16-AS1 and HK2 mRNA competitive binding IGF2BP3.Conclusions1.The expression of MYO16-AS1 in LUAD tissues was significantly down-regulated.2.In vitro experiments found that overexpressed MYO16-AS1 inhibited the invasion and migration of LUAD cells;In vivo,it was found that overexpression of MYO16-AS1 inhibited the tumorigenic ability of LUAD.3.In the cytoplasm,MYO16-AS1 forms a complex with IGF2BP3,which reduces the binding efficiency of IGF2BP3 and HK2 mRNA,inhibits the role of IGF2BP3 in stabilizing HK2 mRNA,and then down-regulates the expression level of HK2 protein and inhibits the invasion and migration ability of LUAD.