HNRNPC Promotes Tumor Progression And Aerobic Glycolysis In Lung Adenocarcinoma Via Up-regulating PKM2 By Affecting PKM Alternative Splicing

Objective: The hnRNPs are a family of broadly functional RNA-binding proteins that bind to pre-m RNA and form ribonucleoprotein particles that perform functions such as transcript stabilization,splicing regulation,polyadenylation,and cytosolic export.The proteins in hnRNP family play an important role in tumor cell apoptosis,angiogenesis,cell invasion and epithelial-mesenchymal transition.Aerobic glycolysis is a key feature of tumor metabolism,providing energy support for tumor growth as well as important intermediate metabolites required for substance synthesis.The abnormal aerobic glycolysis metabolic pattern not only participates in the proliferation of the tumor,but also reduces the ph of the tumor microenvironment by producing a large amount of lactic acid,inhibits immune infiltration and promotes tumor-associated angiogenesis.As a member of the hnRNP protein family,HNRNPC is involved in the regulation of RNA splicing and stability,and its expression is up-regulated in a variety of tumors and correlated with prognosis,but its mechanism related to affecting tumor progression and regulation of tumor aerobic glycolysis in lung adenocarcinoma is not yet fully defined.Methods: Bioinformatics analysis of the HNRNP family genes in the lung adenocarcinoma dataset from TCGA database was used to screen and identify HNRNPC as the key prognostic gene in lung adenocarcinoma.The clinical diagnostic predictive performance of HNRNPC in lung adenocarcinoma was analyzed by using receiver operating curve(ROC).Statistical analysis was performed by immunohistochemical staining scores and clinical information to verify the correlation between HNRNPC expression in protein level and clinical TNM stage.The biological role of HNRNPC in lung adenocarcinoma was verified by functional experiments in vitro and in vitro.Gene set enrichment analysis(GSEA)was used to predict that HNRNPC may regulate aerobic glycolysis in lung adenocarcinoma and the predictions were confirmed by measuring the extracellular acidification rate(ECAR),oxygen consumption rate(OCR),ATP and lactate production in lung adenocarcinoma cells.The regulation of HNRNPC on the expression of glycolysis key genes was examined by Western blot assay and downstream rescue assay was performed by small interfering RNA(si RNA).Finally,RNA sequencing,Southern blot and Western blot were used to explore the specific regulatory mechanisms of HNRNPC on key genes of glycolysis.Results:1.HNRNPC expression was elevated in lung adenocarcinoma tumor tissues,correlated with lung adenocarcinoma-related clinical TNM stage and STAGE grading,and was associated with prognosis independently.2.HNRNPC promotes the ability of lung adenocarcinoma cells to proliferate and migrate in vitro and in vivo.3.HNRNPC promotes aerobic glycolysis in lung adenocarcinoma.4.HNRNPC promotes proliferation,migratory capacity and aerobic glycolysis of lung adenocarcinoma cells in vitro through upregulation of PKM2 expression.5.HNRNPC upregulates PKM2 by alternative splicing of PKM.Conclusion: HNRNPC expression is upregulated in lung adenocarcinoma,and high expression of HNRNPC predicts poor survival outcome.HNRNPC regulates the PKM1/PKM2 expression ratio by regulating alternative splicing of PKM,and upregulates PKM2,leading to proliferation of lung adenocarcinoma cells with enhanced migration ability and promoting aerobic glycolysis in lung adenocarcinoma cells...