Ccdc152 Genes Regulating Tumor Warburg Effect Mechanism Research

Cancer is one of the most common malignant diseases threaten human health.The incidence of cancer has been rising steadily. The study of the pathogenesis and its mechanisms is pivotal for the prevention and treatment of cancer. In recent years, the study of the relationship between cancer metabolism and cancer migration and proliferation has provided new targets for the development of anti-cancer drugs.Tumors exhibit a high glycolytic rate even in the presence of oxygen. This metabolic reprogramming, known as the Warburg effect, is a hallmark of tumor cells. Here, we show that a gene of unknown function, CCDC152(coiled-coil domain containing152), promotes the Warburg effect in tumor cells in a coding-independent manner. Mechanistically, CCDC1523’-UTR assembles as a tail-to-tail natural antisense transcript (NAT) to selenoprotein-P (SEPP1)3’-UTR and suppresses SEPP1translation. Suppression of SEPP1results in the increased phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), the redistribution of selenium (Se) among the cellular selenoproteins, and collectively, enhanced aerobic glycolysis and lipid accumulation. Finally, we find that CCDC152transcription is downregulated by tumor suppressor gene p53and upregulated by transforming growth factor-β1(TGF-β1).MATERIALS AND METHODS:1)To study the mechanism of CCDC152regulating the expression of SEPP1: Firstly,transcription vectors for human SEPP1, CCDC152have been constructed.RNase Protection Assay(RPA) was used to prove the NATs structure between CCDC152and Selenoprotein P; Secondly, HepG2cells were transfected with CCDC1523’-UTR or its ORF region, then the expression of SEPP1and Ago2were detected by using Western blot.Finally, HepG2cells transfected with CCDC1523’-UTR or its ORF were analyzed for the expression of cellular selenoproteins by Real-time PCR quantitative PCR.2)To study CCDC1523’-UTR expression on the Warburg effect and lipid accumulation in the HepG2cells:Firstly, cells were transfected with CCDC1523’-UTR or its ORF region, then the expression of AMPKa and P-AMPKa were detected by using Western blot. Secondly, Warburg effect was evaluated by measuring the consumption of glucose and glutamine in the culture medium and the accumulation of cellular lactate. Finally, HepG2cells transfected with CCDC1523’-UTR or its ORF were stained by Oil-Red O reagent.3)To study the regulatory effect of TGF-(3or p53on CCDC152transcription:293T cells were co-transfected with CCDC152promoter-luciferase vector and p53or Smad2/3vectors, and then the promoter activity was measured by luciferase assay;HepG2cells were treated with TGF-β,then,endogenous CCDC152and SEPP1expression were assayed by qPCR.RESULTS:1)RPA proved the NATs structure between CCDC152and SEPP1. SEPP1expression was significantly reduced in the cells transfected with CCDC1523’UTR but not in the vector bearing only the ORF region,as tested by Western blot.Real-time PCR results showed that suppression of SEPP1resulted in a significant elevated expression of deiodinase-1(DIO1).2)HepG2cells transfected with CCDC1523’-UTR showed enhanced aerobic glycolysis (increased glucose and glutamine consumption and lactate production).The results showed that AMPK phosphorylation(Thr172) was increased in CCDC1523’-UTR transfected cells.3)We identified one putative p53and two Smad binding sites in the CCDC152promoter.The results demonstrated that TGF-β treatment or Smad2/3co-transfection activates the CCDC152transription,whereas p53shows an inhibitory effect.CONCLUSIONS:1)CCDC1523’UTR assembles as a tail-to-tail NAT to SEPP13’-UTR and suppresses SEPP13’-UTR and suppresses SEPP1translation.2)Suppression of SEPP1results in the increased phosphorylation of AMPK, the redistribution of selenium (Se) among the cellular selenoproteins, with a significant elevation of DIO1expression, and collectively, enhanced aerobic glycolysis and lipid accumulation.Find that CCDC152transcription was downregulated by tumor suppressor gene p53and upregulated by TGF-β1.Our study suggests that CCDC152may be an oncogene and a therapeutic target for cancer and liver steatosis.